PowerWaveX Operator`s Manual - Bio-Tek

PowerWaveX Operator`s Manual - Bio-Tek
Microplate Scanning Spectrophotometer
Operator’s Manual
.
7261014 Rev D2
July 2005
Page 1 of 2
This Manual Update contains changes to the PowerWaveX™ Operator’s Manual.
These changes will be incorporated in the next full revision (Rev E) of the Manual.
Page 4-17, Liquid Test 1
Recommendation:
After pipetting the diluted test solution into the microplate (step 3) and
before reading the plate, we strongly recommend shaking the plate at
Variable speed for four minutes. This will allow any air bubbles in the
solution to settle and the meniscus to stabilize. Alternatively, wait 20
minutes after pipetting the diluted test solution before reading the plate.
Pages 4-7 through 4-13, Universal Plate Test
Clarification:
Bio-Tek’s “Absorbance Test Plate” (part number 7260522) can be
used to test the alignment, accuracy, repeatability, and linearity of your
PowerWaveX Microplate Spectrophotometer. Revision D of the Operator’s
Manual refers to this test plate by its former name, the “Universal Test
Plate.”
BIO-TEK INSTRUMENTS, INC., P.O. BOX 998, HIGHLAND PARK, WINOOSKI, VERMONT 05404 USA
COPYRIGHT © 2005 TEL: (888) 451-5171 FAX: (802) 655-7941 Outside the USA: (802) 655-4740 E-mail: [email protected]
www.biotek.com
Creating Solutions for the Future of Science™
7261014 Rev D2
July 2005
Page 2 of 2
Page 4-2, Recommended Test Schedule
Correction:
The schedule shown in Table 4-1 defines the factory-recommended intervals
for performance testing for a microplate reader used for one shift seven days
a week. Note: The risk factors associated with your tests may require that the
Operational and Performance Qualification procedures be performed more
or less frequently than shown below.
Table 4-1 Recommended Test Schedule
Installation
Qualification
Operational
Qualification
Performance
Qualification
Performance
Qualification
Initially & Annually
Monthly
Quarterly
System Self-Test, p. 4-4
9
9
9
Absorbance Plate Test,
p. 4-7
9
9
9
Liquid Test 1*, p. 4-17 or
Liquid Test 2*, p. 4-19
9
Liquid Test 3**, p. 4-22
9
Robotic Lube, p. 4-25
9
Every six months, or after 10,000 cycles.
* If you have an Absorbance Test Plate, run Liquid Test 1. If you do not have an
Absorbance Test Plate, run Liquid Test 2.
** Liquid Test 3 is optional; it is provided for sites requiring verification at wavelengths
lower than those attainable with the Absorbance Test Plate.
Liquid Tests should be performed as part of the Operational Qualification,
during initial installation and annually, as shown in this revised chart.
BIO-TEK INSTRUMENTS, INC., P.O. BOX 998, HIGHLAND PARK, WINOOSKI, VERMONT 05404 USA
COPYRIGHT © 2005 TEL: (888) 451-5171 FAX: (802) 655-7941 Outside the USA: (802) 655-4740 E-mail: [email protected]
www.biotek.com
Creating Solutions for the Future of Science™
PowerWaveX
TM
Microplate Scanning Spectrophotometer
Operator's Manual
For R & D Use
and
in vitro Diagnostic Use
OCTOBER 2002
COPYRIGHT  2002
PART NUMBER 7261014
REVISION D

BIO-TEK INSTRUMENTS, INC.
ii
Preface
Notices
®
BIO-TEK INSTRUMENTS, INC.
Highland Park, Box 998
Winooski, Vermont, 05404-0998
USA
Customer Service and Sales
Internet:
Phone:
Fax:
E-Mail:
www.biotek.com
888-451-5171 (toll free in the U.S.)
802-655-4040 (outside the U.S.)
802-655-7941
[email protected]
Service/TAC
Phone:
Fax:
E-Mail:
800-242-4685 (toll free in the U.S.)
802-655-4740 (outside the U.S.)
802-655-3399
[email protected]
European Coordination Center
Bio-Tek® Instruments GmbH
Kocherwaldstr. 34
D-74177 Bad Friedrichshall
Germany
Phone:
Fax:
E-Mail:
+49 (0) 7136-9680
+49 (0) 7136-968-111
[email protected]
COPYRIGHT
Copyright © 2002, Bio-Tek® Instruments, Incorporated. No part of this publication may be
reproduced, transmitted, transcribed, stored in a retrieval system, or translated into any language
without the written permission of Bio-Tek® Instruments, Incorporated.
TRADEMARKS
Bio-Tek is a registered trademark, and PowerWave X™, KC4™, and KCjunior™ are trademarks of
Bio-Tek Instruments, Inc. Bio-Cell™ is a trademark of Bio-Tek Instruments, and Bio-Cell is
patented under U.S. patent number 5,963,318.
Microsoft is a registered trademark, and Windows, Windows 95/98/2000/NT and XP, Word and
Excel are registered trademarks of Microsoft Corporation.
All other trademarks are the property of their respective holders.
PowerWaveX Operator’s Manual
iii
RESTRICTIONS AND LIABILITIES
Information in this document is subject to change, and does not represent a commitment by
Bio-Tek® Instruments, Inc. Changes made to the information in this document will be incorporated
in new editions of the publication. No responsibility is assumed by Bio-Tek for the use or
reliability of software or equipment that is not supplied by Bio-Tek, or its affiliated dealers.
REVISION HISTORY
Revision
Date
Change
A
8/98
First Release.
B
2/99
Incorporated Manual Updates.
C
8/99
Added support for PowerWavex Select model.
Enhanced Chapter 4, Performance Verification & Qualification. Assigned each
of the recommended maintenance tasks the Installation, Performance, and/or
Operation Qualification classification.
D
10/02
Changed PowerWavex to PowerWaveX throughout. Updated Notices, Chapters 1
through 4, and Appendices B and C. Incorporated manual update for periodic
mantenance of robotic units in Ch 4. Added Cleaning section to Appendix A.
DOCUMENT CONVENTIONS
This manual uses the following typographic conventions.
Example
!
iv
Description
This icon calls attention to important safety notes.
Warning!
A Warning indicates the potential for bodily harm and tells you how to
avoid the problem.
Caution:
A Caution indicates potential damage to the instrument and tells you how
to avoid the problem.
DEFINE
Text in COURIER font represents menu options as they appear in the
instrument’s display.
Note:
Bold text is primarily used for emphasis.
iii
This icon calls attention to important information.
Preface
Warnings
!
The PowerWaveX should be operated on a flat surface away from direct sunlight or
strong incandescent light. Excessive humidity should be avoided.
Please read the following hazards and cautions before operating the instrument!
HAZARDS_________________________________________________________________
Warning! Power Rating. The PowerWaveX must be connected to a power receptacle that provides
voltage and current within the specified rating for the system. Use of an incompatible power
receptacle may produce electrical shock and fire hazards.
Warning! Internal Voltage. Always turn off the power switch and unplug the power cord before
cleaning the instrument’s outer surface.
Warning! Liquids. Avoid spilling liquids on the reader; fluid seepage into internal components
creates a potential shock hazard. Do not operate the instrument if internal components are exposed
to fluid.
Warning! Software. The microplate reader operator must follow the manufacturer’s assay package
insert when modifying software parameters and establishing result calculation methods, using the
reader’s on-board software.
Warning! Data Reduction Protocol. The reader’s software will flag properly defined controls
when they are out of range. It will present all the data with the appropriate error flags in order for
the user to determine their validity. Because there have been no limits applied to the raw
absorbance data, all information exported via computer control must be analyzed completely.
Warning! Unspecified Use. Failure to operate this equipment according to the guidelines and
safeguards specified in this manual could result in a hazardous condition.
PRECAUTIONS _____________________________________________________________
The following precautions are provided to help you avoid damaging the system:
Caution: Service. The system should be serviced by authorized service personnel. Only qualified
technical personnel should perform troubleshooting and service procedures on internal components.
Caution: Environmental Conditions. Do not expose the system to temperature extremes.
Ambient temperatures should remain between 18-40°C. System performance may be adversely
affected if temperatures fluctuate above or below this range.
PowerWaveX Operator’s Manual
v
Caution: Sodium Hypochlorite. Do not expose any part of the instrument to Sodium Hypochlorite
solution (bleach) for more than 30 minutes. Prolonged contact may damage the instrument surfaces.
Be certain to rinse and thoroughly wipe all surfaces.
Caution: Power Supply. Only use the correct line voltage when operating the Automated
Microplate Reader. A four-position line voltage select switch is used to adjust for different line
voltages. This switch is located on the power input module. See the section Adjusting Line Voltage
Input Range in Chapter 2 for more details.
Caution: Carrier Retention Screw. The carrier retention screw must be removed prior to operating
the device.
vi
Preface
Electromagnetic Interference and Susceptibility
USA FCC CLASS A
Warning: Changes or modifications to this unit not expressly approved by the manufacturer could
void the user's authority to operate the equipment.
This equipment has been tested and found to comply with the limits for a Class A digital device,
pursuant to Part 15 of the FCC Rules.
These limits are designed to provide reasonable protection against harmful interference when the
equipment is operated in a commercial environment. Like all similar equipment, this equipment
generates, uses, and can radiate radio frequency energy and, if not installed and used in accordance
with the instruction manual, may cause harmful interference to radio communications. Operation of
this equipment in a residential area is likely to cause interference, in which case the user will be
required to correct the interference at his own expense.
Canadian Department of Communications Class A
This digital apparatus does not exceed Class A limits for radio emissions from digital apparatus set
out in the Radio Interference Regulations of the Canadian Department of Communications.
Le present appareil numerique n'met pas du bruits radioelectriques depassant les limites applicables
aux appareils numerique de la Class A prescrites dans le Reglement sur le brouillage radioelectrique
edicte par le ministere des Communications du Canada.
User Safety
This device has been type tested by an independent laboratory and found to meet the requirements
of the following:
•
Canadian Standards Association CAN/CSA C22.2 No. 1010.1-1992
"Safety Requirements for Electrical Equipment for Measurement, Control
and Laboratory Use, Part 1: General Requirements".
PowerWaveX Operator’s Manual
vii
Based on the testing described below, this instrument bears the CE mark.
EMC EC DIRECTIVE 89/336/EEC
•
EN 50081-1, CLASS A-Emissions
The system has been type tested by an independent testing laboratory and found to meet the
requirements of EC Directive 89/336/EEC for Radiated Emissions and Line Conducted
Emissions. Verification was to the limits and methods of EN 55022. The device is classified
as EN 55022, Class A.
•
EN 50082-1 Immunity
The system was also tested and found to meet requirements for Electrostatic Discharge
Susceptibility, Radiated Susceptibility, and Electrical Fast Transient/Burst Susceptibility.
Verification of compliance was conducted to the limits and methods of EN 50082-1:1992;
IEC 1000-4-2:1995; IEC 1000-4-3:1995; IEC 1000-4-4:1995; EN 61000-4-6:1996;
EN 61000-4-11: 1994.
EC Directive 73/23/EEC Low Voltage (Safety)
The system has been type tested by an independent testing laboratory and was found to meet the
requirements of EC Directive 73/23/EEC for Low Voltage. Verification of compliance was
conducted to the limits and methods of the following:
•
EN 61010-1: 1993
"Safety requirement for electrical equipment for measurement, control and laboratory use.
Part 1, General requirements" (including amendment No. 2: 1995).
viii
Preface
Safety Symbols
The following warning and informational symbols may be found in various locations on the
PowerWaveX. Only qualified personnel who recognize shock hazards and are familiar with the
safety precautions should use this instrument. Read the manual carefully before operating this
instrument.
Alternating current
Courant alternatif
Dreiphasen-Wechselstrom
Corriente Alterna
Corrente alternata
Earth ground terminal
Borne de terre
Erde (Bettriebserde)
Borne de Tierra
Terra (di funzionamento)
Protective conductor terminal
Borne de terre de protection
Schutzleiteranscluss
Borne de Tierra de Protección
Terra di protezione
On (Supply)
Marche (alimentation)
Ein (Verbindung mit dem Netz)
Conectado
Chiuso
Off (Supply)
Arrêt (alimentation)
Aus (Trennung vom Netz)
Desconectado
Aperto (sconnessione dalla rete di alimentazione)
PowerWaveX Operator’s Manual
ix
Caution, risk of electric shock
Attention, risque de choc electrique
Gefährliche elektrische Spannung
Atención, riesgo de sacudida eléctrica
Alta tensione (in questo documento Alta tensione non significa
“tensione pericolosa” come definito in IEC 417)
Caution (refer to accompanying documents)
Attention (voir documents d'accompanement)
Achtung siehe Begleitpapiere
Atención (vease los documentos incluidos)
Attenzione, consultare la doc annessa
x
Preface
Intended Use Statement
The PowerWaveX is an eight-channel, automated, benchtop, general-purpose, Microplate
Spectrophotometer that performs in vitro diagnostic analyses of a variety of samples. The
Performance Characteristics of the data reduction software have not been established with any
laboratory diagnostic assay. The user must evaluate this software in conjunction with the specific
laboratory diagnostic assay. This evaluation must include the confirmation that new performance
characteristics for the specific assay are met.
This system is designed for use with a variety of microplate-based assays. A versatile curve-fitting
and statistical software program is preloaded on every PowerWaveX; plate templates and formulas
are automatically combined with the protocol assay setup. Data results may be printed out, or sent
to a computer running a Bio-Tek software package, such as KCjunior or KC4 for Windows®.
The on-board software provides:
•
An easy-to-use, menu-driven interface
•
Endpoint curvilinear regressional and statistical calculations
•
Curve fitting, with 4-parameter, cubic, quadratic, linear, cubic-spline, point-to-point
and 2-P (Logit) methods
•
Formula calculations for more complex mathematical operations
•
Ability to define controls and positive and negative cutoffs
•
Kinetic read capability, with maximum slope, R-squared at maximum slope and Onset
OD Time analysis
•
Ability to perform a spectral scan reading from 200 to 999 nm or
340 to 999 nm, depending on the model
•
Scanning read mode to provide area under the curve calculations
•
Optional preprogrammed common biological assays
Specimen Preparation
Samples should be obtained, treated, and stored following instructions and recommendations
determined by the user’s laboratory.
Quality Control
It is considered good laboratory practice to run laboratory samples according to instructions and
specific recommendations included in the package insert or standard laboratory protocol for the test
to be conducted. Failure to conduct Quality Control checks could result in erroneous test data.
PowerWaveX Operator’s Manual
xi
About This Manual
The intent of this Operator’s Manual is to quickly instruct the new user how to set up and operate
Bio-Tek’s PowerWaveX. To help you read and understand this manual, certain document
conventions have been used.
Major topic headings start a new page (such as About This Manual, above) to give you a visual
and style clue that a new major subject is being introduced. One or more subheadings may appear
below each major heading.
Registration Card
Once the PowerWaveX has been set up and is running successfully, please take a moment
to fill out and mail the postage-paid Warranty Registration card. By sending in the registration
card, you will be assured of receiving prompt information on product enhancements.
xii
Preface
Warranty
This Warranty is limited and applies only to new products, except for computer-based software,
which is covered under a separate Warranty Policy, manufactured by Bio-Tek Instruments, Inc.
(“Bio-Tek”). Bio-Tek makes no warranty whatsoever regarding the condition of used products.
Bio-Tek warrants the instrument (hereinafter collectively referred to as “Products” or “Product”) for
a period of one (1) year from the original purchase date against defective materials or workmanship.
This Warranty is limited to the original purchaser (the “Purchaser”) and cannot be assigned or
transferred. All claims under this Limited Warranty must be made in writing to Bio-Tek, Attention:
Service Department. Purchaser must ship the Product to Bio-Tek, postage pre-paid. Bio-Tek shall
either repair or replace with new or like new, at its option and without cost to the Purchaser, any
Product which in Bio-Tek’s sole judgment is defective by reason of defects in the materials or
workmanship.
This Warranty is VOID if the Product has been damaged by accident or misuse, or has been
damaged by abuse or negligence in the operation or maintenance of the Product, including without
limitation unsafe operation, operation by untrained personnel, and failure to perform routine
maintenance. This Warranty is VOID if the Product has been repaired or altered by persons not
authorized by Bio-Tek, or if the Product has had the serial number altered, effaced, or removed.
This Warranty is VOID if any of the Products has not been connected, installed or adjusted strictly
in accordance with written directions furnished by Bio-Tek. Batteries, fuses, light bulbs, and other
“consumable” items used in any of the Products are not covered by this Warranty. Software utilized
in conjunction with any of the Products is not covered by the terms of this Warranty but may be
covered under a separate Bio-Tek software warranty.
We will continue to stock parts for a maximum period of five (5) years after the manufacture of any
equipment has been discontinued. Parts shall include all materials, charts, instructions, diagrams,
and accessories that were furnished with the standard models.
THIS WARRANTY CONTAINS THE ENTIRE OBLIGATION OF BIO-TEK INSTRUMENTS,
INC., AND NO OTHER WARRANTIES, EXPRESSED, IMPLIED, OR STATUTORY ARE
GIVEN. PURCHASER AGREES TO ASSUME ALL LIABILITY FOR ANY DAMAGES
AND/OR BODILY INJURY OR DEATH THAT MAY RESULT FROM THE USE OR MISUSE
OF ANY EQUIPMENT OR INSTRUMENT BY THE PURCHASER, HIS EMPLOYEES,
AGENTS, OR CUSTOMERS, OTHER THAN THE EXPRESS WARRANTY CONTAINED
HEREIN. WE SHALL NOT BE RESPONSIBLE FOR ANY DIRECT OR CONSEQUENTIAL
DAMAGES OF ANY KIND. THIS WARRANTY SHALL NOT BE CHANGED OR MODIFIED
IN ANY WAY WITHOUT THE EXPRESS WRITTEN PERMISSION OF AN OFFICER OF
BIO-TEK INSTRUMENTS, INC.
THIS WARRANTY IS VOID UNLESS THE PURCHASE REGISTRATION CARD HAS BEEN
COMPLETED AND MAILED TO US WITHIN TEN (10) DAYS OF PURCHASE.
PowerWaveX Operator’s Manual
xiii
xiv
Preface
Table of Contents
Preface...........................................................................................................iii
Notices ............................................................................................................................... iii
Copyright ..................................................................................................................... iii
Trademarks ................................................................................................................. iii
Restrictions and Liabilities ........................................................................................... iv
Revision History........................................................................................................... iv
Document Conventions ............................................................................................... iv
Warnings and Precautions...................................................................................................v
Hazard Warnings..........................................................................................................v
Precautions ..................................................................................................................v
Electromagnetic Interference and Susceptibility ................................................................ vii
CE Mark Information......................................................................................................... viii
Safety Symbols................................................................................................................... ix
Intended Use Statement ..................................................................................................... xi
Specimen Preparation ................................................................................................. xi
Quality Control............................................................................................................. xi
About this Manual.............................................................................................................. xii
Registration Card............................................................................................................... xii
Warranty ........................................................................................................................... xiii
List of Tables and Figures ................................................................................................ xxi
Chapter 1 Introduction ...............................................................................1-1
Introducing the PowerWaveX .......................................................................................... 1-1
Variations ................................................................................................................. 1-1
Hardware Features .......................................................................................................... 1-2
Software Features ........................................................................................................... 1-2
Package Contents ........................................................................................................... 1-3
Optional Accessories ....................................................................................................... 1-3
Specifications .................................................................................................................. 1-4
Technical Support............................................................................................................ 1-8
Chapter 2: Installation ................................................................................2-1
Operating Environment.................................................................................................... 2-1
Unpacking and Repackaging the Instrument ................................................................... 2-1
PowerWaveX Operator’s Manual
xv
Removal of the Carrier Retention Screw in PowerWaveX Select Models........................ 2-3
After Unpacking, Verify Performance ....................................................................... 2-4
Before Repackaging the Instrument ......................................................................... 2-4
Adjusting the Reader’s Wavelength Table Settings ......................................................... 2-4
Printing and Data Communications ................................................................................. 2-6
Installing a Printer..................................................................................................... 2-6
Parallel Port Pin Definition........................................................................................ 2-7
Serial Port for Communication with Other Devices .................................................. 2-8
Connecting to the PC and Establishing Communication .......................................... 2-8
Serial Port Pinout Description................................................................................... 2-8
Setting Up Communications Parameters.................................................................. 2-9
Components .................................................................................................................. 2-11
Lamp ...................................................................................................................... 2-11
Cooling Fan ............................................................................................................ 2-11
Adjusting the Line Voltage Input Range......................................................................... 2-12
Adjusting the Fuse Configuration or Fuse Replacement................................................ 2-14
Other Utility Options ...................................................................................................... 2-15
Setting the Date and Time...................................................................................... 2-16
Specifying Data Output and Reporting Options ...................................................... 2-17
Data Output..................................................................................................... 2-17
Compatible Printers......................................................................................... 2-17
Report Types................................................................................................... 2-18
Selecting Read Options.......................................................................................... 2-19
Chapter 3: Operation ..................................................................................3-1
Overview.......................................................................................................................... 3-1
Front Panel ...................................................................................................................... 3-2
Selecting Menu Functions ........................................................................................ 3-2
Scrolling Through Options ........................................................................................ 3-2
Entering Assay Names ............................................................................................. 3-3
Moving Backward Through the Menus ..................................................................... 3-3
Saving Assays.......................................................................................................... 3-3
Starting or Stopping a Read ..................................................................................... 3-3
Startup Screen................................................................................................................. 3-3
Main Menu....................................................................................................................... 3-4
DEFINE ........................................................................................................................... 3-5
Selecting an Assay ................................................................................................... 3-6
Editing the Assay Name ........................................................................................... 3-7
xvi
Preface
Define METHOD.............................................................................................................. 3-8
Read Type................................................................................................................ 3-8
Spectral Scanning ............................................................................................. 3-9
Delay First Read..................................................................................................... 3-10
Incubation Temperature (Reader-Dependent) ....................................................... 3-10
Wavelength(s) ........................................................................................................ 3-11
Measurement Wavelength(s) ................................................................................. 3-11
Kinetic Read Parameters........................................................................................ 3-12
Kinetic Interval ................................................................................................ 3-12
Total Number of Kinetic Reads ....................................................................... 3-13
Kinetic Duration ............................................................................................... 3-13
Plate Shaking ......................................................................................................... 3-13
Shake Time ..................................................................................................... 3-14
Shake Speed................................................................................................... 3-14
Kinetic Analysis ...................................................................................................... 3-15
Kinetic Points .................................................................................................. 3-15
Onset OD ........................................................................................................ 3-16
Well Scanning ........................................................................................................ 3-16
Define MAP ................................................................................................................... 3-17
Map Generation...................................................................................................... 3-18
Mapping Direction ........................................................................................... 3-18
Replication Direction ....................................................................................... 3-19
Start Mapping at Well Location........................................................................ 3-21
Blank Map ....................................................................................................... 3-21
Blanking Constant ........................................................................................... 3-22
Number of Blanks............................................................................................ 3-22
Blank Location................................................................................................. 3-23
Number of Standards ...................................................................................... 3-23
Number of Standard Replicates ...................................................................... 3-24
Average Standards.......................................................................................... 3-24
Standard Concentrations................................................................................. 3-25
Number of Controls ......................................................................................... 3-25
Control Type.................................................................................................... 3-26
Number of Control Replicates ......................................................................... 3-26
Location of Controls ........................................................................................ 3-27
Number of Samples......................................................................................... 3-27
Number of Sample Replicates......................................................................... 3-27
Sample Location.............................................................................................. 3-28
PowerWaveX Operator’s Manual
xvii
Define FORMULA.......................................................................................................... 3-29
Calculation Structure .............................................................................................. 3-29
Formula Type ......................................................................................................... 3-30
Formula Entry......................................................................................................... 3-31
MATH .............................................................................................................. 3-32
OTHER............................................................................................................ 3-32
MAP ................................................................................................................ 3-32
FUNCTION...................................................................................................... 3-32
Validation Formulas................................................................................................ 3-33
Control and Blank Validation Formulas ........................................................... 3-33
Examples ................................................................................................. 3-33
Number of Required Blanks/Controls .............................................................. 3-34
Assay Validation Formulas .............................................................................. 3-34
Examples ................................................................................................. 3-34
Transformation Formulas ....................................................................................... 3-35
Simple Transformation Formulas .................................................................... 3-35
Transformation Variable (TVAR) ..................................................................... 3-35
Examples ........................................................................................................ 3-36
Cutoff Formulas...................................................................................................... 3-38
Greyzone......................................................................................................... 3-38
POS/NEG Calls ............................................................................................... 3-39
Examples ........................................................................................................ 3-39
General Formulas................................................................................................... 3-40
Define CURVE............................................................................................................... 3-41
Curve-Fit Type ................................................................................................ 3-41
Edit Standard Outliers ..................................................................................... 3-42
X/Y Axis Type.................................................................................................. 3-43
Extrapolation of Unknowns.............................................................................. 3-43
Panel Assays................................................................................................................. 3-44
Reading a Microplate..................................................................................................... 3-47
Selecting an Assay ................................................................................................. 3-48
Run-Time Prompts ................................................................................................. 3-48
Enter Number of Samples ............................................................................... 3-49
Enter Plate ID.................................................................................................. 3-49
Enter Sample ID .............................................................................................. 3-50
Enter Well Location ......................................................................................... 3-50
Beginning the Plate Read....................................................................................... 3-51
xviii
Preface
Printing Reports............................................................................................................. 3-52
Results Report ................................................................................................ 3-53
Editing Standard Outliers ......................................................................... 3-53
Printing Results ........................................................................................ 3-54
Map Report ..................................................................................................... 3-55
Assay Report................................................................................................... 3-55
List Report....................................................................................................... 3-55
Chapter 4: Performance Verification/Qualification Tests ......................4-1
Recommendations for Achieving Optimum Performance ................................................ 4-1
Recommended Test Schedule ........................................................................................ 4-2
Installation and Performance Qualification Procedures ................................................... 4-2
Routine Procedure........................................................................................................... 4-3
System Test ............................................................................................................. 4-4
Photodiodes ...................................................................................................... 4-4
Xenon Flash Lamp ............................................................................................ 4-4
Incubation.......................................................................................................... 4-4
Checksum Test (CHKSUM)...................................................................................... 4-6
Calibration Verification..................................................................................................... 4-7
Universal Plate Test ................................................................................................. 4-7
Requirements .................................................................................................... 4-8
Entering the Universal Test Plate Data ............................................................. 4-8
Running the Universal Plate Test ...................................................................... 4-9
Wavelength Accuracy...................................................................................... 4-13
Liquid Testing ................................................................................................................ 4-14
Stock Solution Formulation..................................................................................... 4-15
Procedure A .................................................................................................... 4-15
Procedure B .................................................................................................... 4-16
Liquid Test 1........................................................................................................... 4-17
Liquid Test 2........................................................................................................... 4-19
Linearity – Test A ............................................................................................ 4-20
Repeatability – Test B ..................................................................................... 4-20
Channel-to-Channel Variation and Alignment – Test C ................................... 4-21
Liquid Test 3........................................................................................................... 4-22
Procedure A .................................................................................................... 4-23
Procedure B .................................................................................................... 4-23
Liquid Test 3 Procedure .................................................................................. 4-23
PowerWaveX Operator’s Manual
xix
Repeatability – Test A ..................................................................................... 4-24
Linearity – Test B ............................................................................................ 4-24
Maintenance of Robotic Units........................................................................................ 4-25
Appendix A: Decontamination and Cleaning.......................................... A-1
Decontamination Procedure ............................................................................................ A-1
Purpose .................................................................................................................... A-1
General Considerations............................................................................................ A-1
Procedure................................................................................................................. A-2
Cleaning Procedure ......................................................................................................... A-2
Appendix B: Computer Control................................................................ B-1
Overview.......................................................................................................................... B-1
Controlling the Reader with KC4...................................................................................... B-2
Problems .................................................................................................................. B-2
Getting Started with KC4 .......................................................................................... B-3
Controlling the Reader with KCjunior............................................................................... B-4
Problems .................................................................................................................. B-4
Getting Started with KCjunior ................................................................................... B-5
Controlling the Reader Using Serial Protocol .................................................................. B-6
Computer Control Command Set .................................................................................... B-6
Using the Stop Key to Halt Plate Scans and Reads ...................................................... B-20
Status String Format...................................................................................................... B-21
Appendix C: Error Codes.......................................................................... C-1
Appendix D: Sample Reports ................................................................... D-1
xx
Preface
List of Ilustrations
Figure 2-1:
Unpacking and Repackaging the Reader ................................................................... 2-2
Figure 2-2:
Removal of the Carrier Retention Screw and Accompanying Washers from
the PowerWaveX Select .............................................................................................. 2-3
Figure 2-3:
Serial and Parallel Connectors .................................................................................... 2-6
Figure 2-4A:
Voltage Selector Board and Fuse Holder.................................................................. 2-12
Figure 2-4B:
Selecting Fuse Position ............................................................................................. 2-12
Figure 2-5:
Options Available Under UTIL ................................................................................ 2-15
Figure 3-1:
PowerWaveX Front Panel ........................................................................................... 3-2
Figure 3-2:
Options Available From the Main Menu .................................................................... 3-4
Figure 3-3:
Options Available Under DEFINE ............................................................................. 3-5
Figure 3-4:
Examples of Mapping Directions............................................................................. 3-20
Figure 3-5:
Options Available Under READ............................................................................... 3-47
Figure 4-1:
Sample Output for the System Test............................................................................. 4-5
Figure 4-2:
Sample Universal Test Plate Data Sheet ..................................................................... 4-8
Figure 4-3:
Sample Printout Showing the Calibration Plate Analysis ......................................... 4-11
Figure 4-4:
Sample Data Sheet Showing Wavelength of Peak in the Interval Between
580 and 590 nm......................................................................................................... 4-13
Figure 4-5:
Location of Bracket, Motor Shaft, Mylar Sheet, and Main PCB/Motor Assembly .. 4-27
Figure 4-6:
Roller Surface and Hook-In Bracket......................................................................... 4-28
Figure D-1:
Matrix Report Showing Well Ids, Calculated OD Values and Concentrations,
Calls on Samples, and Comments ...............................................................................D-2
Figure D-2:
Curve Fit Report Showing the Curve Method, Equation, Coefficients, and
R-Squared Value .........................................................................................................D-3
Figure D-3:
First Page of Column Report, Showing Plate Data for Blanks and Standards,
the Interpretation of Results, and Comments ..............................................................D-4
Figure D-4:
Second Page of Column Report, Showing Plate Data for Samples ............................D-5
Figure D-5:
Column Report Without Samples ...............................................................................D-6
Figure D-6:
Panel Report................................................................................................................D-7
Figure D-7:
Assay Detail Report ....................................................................................................D-8
Figure D-8
Assay List....................................................................................................................D-9
PowerWaveX Operator’s Manual
xxi
List of Tables
xxii
Table 2-1:
Parallel Connector Pinouts................................................................................... 2-7
Table 2-2:
Serial Pinout Description ..................................................................................... 2-8
Table 4-1:
Recommended Test Schedule .............................................................................. 4-2
Table 4-2:
Typical Enzyme-Substrate Combinations and Stopping Solutions.................... 4-14
Table 4-3:
Stock Solution Formulation for Liquid Test Nos. 1 and 2................................. 4-15
Table 4-4:
Test Tube Dilutions............................................................................................ 4-19
Table 4-5:
Phosphate Buffered Saline 10X Concentrate Solution ...................................... 4-22
Table B-1:
Serial Cable Pin-Out Description.........................................................................B-3
Table B-2:
ASCII Control Characters Used in Computer Control Protocol ..........................B-6
Preface
Chapter 1
Introduction
This chapter introduces the PowerWaveX and describes its
hardware and software features and technical
specifications. Instructions on how to contact Bio-Tek for
Technical Assistance are included on page 1-8.
Introducing the PowerWaveX
Bio-Tek’s PowerWaveX is an eight-channel reader-assay system for research and development
and in vitro diagnostic use. The reader can serve as a stand-alone system, or as an integral part of
a PC-based system, using Bio-Tek’s KC4 or KCjunior software packages.
The PowerWaveX is a spectrophotometer that has the capability of spectral scanning using a
monochromator. This allows the user to perform endpoint and kinetic analysis, in 96-well
microplates, using any wavelength between 200 and 999 nm, or 340 and 999 (depending on the
model), thereby achieving the maximum absorbance for any sample.
The reader features superior optical specifications, with an extended dynamic range of up to 4.000
absorbance units.
The instrument’s on-board processor, 2 X 24 character LCD screen, and membrane keypad allow
easy definition and management of assay protocols, templates, formulas, and data. Results can be
output in a printed report format, or exported for use in a variety of microplate-based data
manipulation applications.
Variations
Bio-Tek’s PowerWaveX may be configured with all or selected options:
•
PowerWaveX Select models support the reading of 96- and 384-well plates.
•
“I” model instruments have a 4-Zone incubation chamber that controls the
temperature from 4° above ambient to 50°C.
iii
•
“R” model instruments are robotic compatible.
•
An internal bar code scanner option is also available.
Note: 384-well plate reading must be performed through computer control
or with KC4 or KCjunior software. See Appendix B for more information.
Hardware Features
•
Xenon flash light source
•
Eight optics channels, with an additional reference channel
•
Μonochromator with N range of 200 to 999 nm or 340 to 999 nm, depending on
reader model
•
A 2 X 24 character LCD display
•
A membrane keypad with alphanumeric keys
•
Adjustable plate shake frequency and durations
•
Reads 96-well microplates with 0.355" / 9-mm well centers
•
Reads 384-well microplates with 0.177" / 4.5-mm centers (PowerWaveX Select
model only)
•
Operates from 100, 120, 230, or 240 VAC @ 50-60 Hz (with external switching)
•
One serial COM port (25-pin male connector)
•
One parallel port (25-pin female connector)
•
Internal bar code option
•
4-Zone incubation chamber option
•
Robotics interface option
Software Features
•
Easy-to-use menu-driven interface
•
Endpoint, Kinetic and Scanning calculations
•
Curve fitting, with 4-parameter, cubic, quadratic, linear, 2-P, cubic-spline and
point-to-point methods
•
Transformation and formula calculations for more complex mathematical operations,
including validations
•
Up to 55 assays are available on-board (27 open assays, plus preprogrammed
common biological assays)
•
Automatically stores results for the last 8 plates
•
Spectral scanning using on-board software with report of peak OD and wavelength
at peak OD
1-2
Introduction
Package Contents
•
Microplate scanning spectrophotometer
•
Power cord
•
Operator’s Manual and Warranty Registration Card (PN 7261014)
•
Assay Reference Guide, depending on the model (PN 7271006)
•
Dust cover (PN 7342066)
•
Parallel cable (PN 71072)
•
Serial cable (PN 75053)
•
PowerWaveX Select model only: Allen wrench (PN 49329)
Optional Accessories
•
Service Manual for all PowerWaveX models (PN 7261016)
•
Universal Test Plate (PN 7260522)
•
Bio-Cell™ for 1-cm wavelength readings (PN 7272051)
•
Bio-Cell adaptor plate for containing up to eight Bio-Cells (PN 7270512)
•
Installation-Operational-Performance (IQ-OQ-PQ) package (PN 7260219)
PowerWaveX Operator's Manual
1-3
Specifications
•
Microplates
All models accommodate standard 96-well microplates. The Select model also
accommodates 384-well microplates.
iii
Note: Testing is recommended with 384-well plate types to determine compatibility.
The Select was tested with Greiner and Costar 384-well plates, and with Nunc
black-sided clear-bottom 384-well plates, and results meet specifications.
•
Speed of Reading
The actual plate read time is dependent on the method of reading:
In Normal read mode, the reading speed is variable with the absorbance of the samples.
If a sample measurement is less than 1% of the channel blank signal, the reader will
automatically go into an enhanced read mode where longer read intervals are applied for
increased performance.
In Rapid read mode, enhanced readings are not used; therefore, the reading speed is not
variable.
Wavelength choice will also change read times slightly. Each wavelength has a unique
location within the monochromator, and the different locations require varying amounts of
time to position. This effect is more noticeable in dual-wavelength assays.
96-Well Read Timing
Normal Read Mode
Endpoint
Rapid Read Mode
Endpoint
630 nm
630/450 nm
Single
Dual
23 to 31 sec.
35 to 53 sec.
Single
Dual
23 sec.
35 sec.
Kinetics: 14 seconds from A1 to A1 in rapid mode, single wavelength.
384-Well Read Timing
Normal Read Mode
Endpoint
Rapid Read Mode
Endpoint
630 nm
630/450 nm
Single
Dual
41 to 86 sec.
71 to 141 sec.
Single
Dual
37 sec.
64 sec.
Kinetics: 28 seconds from A1 to A1 in rapid mode, single wavelength.
1-4
Introduction
•
•
Optical Specifications
λ range:
200 to 999 nm or 340 to 999 nm (model-dependent)
λ accuracy:
± 2 nm
λ repeatability:
± 0.2 nm
λ bandpass:
5 nm
Optical Performance
96-Well Plate Reading
The following specifications apply to 96-well, flat- or round-bottom plates, single wavelength
endpoint or kinetic (with intervals of at least 20 seconds) readings. Note: For the following
performance, the Gain on the Optics test should be below 6, in normal read mode.
Endpoint or Kinetic with an interval < 20 seconds:
Absorbance
Measurement Range: 0.000 to 4.000 Abs
Accuracy:
0.000 to 2.000 Abs ± 1% ± 0.010 Abs
2.000 to 3.000 Abs ± 3% ± 0.010 Abs
Linearity:
0.000 to 2.000 Abs ± 1%
2.000 to 3.000 Abs ± 3%
Repeatability:
0.000 to 2.000 Abs ± 1% ± 0.005 Abs
2.000 to 3.000 Abs ± 3% ± 0.005 Abs
Spectral Scanning or Kinetics with short intervals:
When read intervals from 8 seconds to 20 seconds are selected, or spectral scanning is
performed, the specifications are as follows:
Absorbance
Measurement Range: 0.000 to 3.000 Abs
Accuracy:
0.000 to 2.000 Abs ± 1% ± 0.010 Abs
2.000 to 2.500 Abs ± 3% ± 0.010 Abs
Linearity:
0.000 to 2.000 Abs ± 1%
2.000 to 2.500 Abs ± 3%
Repeatability:
0.000 to 2.000 Abs ± 1% ± 0.005 Abs
2.000 to 2.500 Abs ± 3% ± 0.005 Abs
PowerWaveX Operator's Manual
1-5
384-Well Plate Reading
The following specifications apply to 384-well, flat- or round-bottom plates, singlewavelength endpoint or kinetic (with intervals of at least 20 seconds) readings.
Note: For the following performance, the Gain on the Optics Test should be below 6,
in normal read mode.
Endpoint or Kinetic with an interval < 20 seconds:
Absorbance
Measurement Range: 0.000 to 4.000 Abs
Accuracy:
0.000 to 2.000 Abs ± 1% ± 0.010 Abs
2.000 to 2.500 Abs ± 3% ± 0.010 Abs
Linearity:
0.000 to 2.000 Abs ± 2%
2.000 to 2.500 Abs ± 3%
Repeatability:
0.000 to 2.000 Abs ± 2% ± 0.010 Abs
2.000 to 2.500 Abs ± 3% ± 0.010 Abs
Spectral Scanning or Kinetics with short intervals:
When read intervals from 32 seconds to 80 seconds are selected, or spectral scanning is
performed, the specifications are as follows:
Absorbance
Measurement Range: 0.000 to 4.000 Abs
Accuracy:
0.000 to 2.000 Abs ± 1% ± 0.010 Abs
2.000 to 2.500 Abs ± 3% ± 0.010 Abs
Linearity:
0.000 to 2.000 Abs ± 2%
2.000 to 2.500 Abs ± 3%
Repeatability:
0.000 to 2.000 Abs ± 2% ± 0.010 Abs
2.000 to 2.500 Abs ± 3% ± 0.010 Abs
1-6
Introduction
•
Hardware and Environmental Specifications
Display:
2 X 24 Character LCD
Light Source:
Xenon flash light source
- 10 W max. average power
- Life: 1 billion flashes
Dimensions:
16.0” deep X 15.5” wide X 9.75” high
40.1 cc deep X 39.3 cc wide X 24.8 cc high
Weight:
40 lb. maximum (18.2 kg)
Environment:
Operational temperature 18-40°C
Humidity:
10% to 85%, non-condensing
Power Consumption:
100 VA
Four Voltage Ranges accommodated by voltage
selection switch:
Incubation option:
Range 1
100 VAC
90-110 VAC, 50-60 Hz
Range 2
115 VAC
103-127 VAC, 50-60 Hz
Range 3
230 VAC
207-253 VAC, 50-60 Hz
Range 4
240 VAC
216-264 VAC, 50-60 Hz
Temperature Control: 4° over ambient to 50°C
Temperature Variation: ± 0.5°C across the plate @ 37°C
(with the plate sealed)
Internal bar code reader
If enabled, the internal bar code reader recognizes a number of
option:
common bar code types during the plate read operation. Specific
information about each bar code type is available from Bio-Tek
Technical Services. The reader’s bar code option is compatible
with the following bar code types:
Robotic interface option:
CODABAR
UPC
CODE 39
EAN
INTERLEAVED 2 of 5
MSI
CODE 11
PLESSEY
CODE 93
CODE 128
The Robotic interface option allows the reader to function with an
autoloading robot. Using computer control commands from a host
PC, the reader’s functions can be controlled in conjunction with
the robotic system. The Robotic interface model can be configured
with all options available for the standard PowerWaveX.
PowerWaveX Operator's Manual
1-7
Technical Support
Bio-Tek’s PowerWaveX is backed by a superior support staff. If the PowerWaveX ever fails to
work perfectly, please contact Bio-Tek's Technical Assistance Center.
Whichever method of contact you choose, please be prepared to provide the following
information:
•
Product name and serial number.
•
The reader's on-board software configuration information. To see this, start at the
reader main menu and press UTIL
TESTS
CHKSUM.
•
The specific steps that produce your problem.
•
Any error codes displayed on the screen (see Appendix C for information on error
codes).
•
A daytime phone number.
•
Your name and company information.
•
A fax number and/or an email address, if available.
•
If you need to return the reader to Bio-Tek for service, contact Bio-Tek for a Return
Materials Authorization (RMA) number, and be sure to repack the reader properly
(see Chapter 2, Installation).
Phone Support
You can telephone the Technical Assistance Center between 8:30 AM and 5:30 PM Eastern
Standard Time (EST), Monday through Friday, excluding holidays.
Bio-Tek Instruments Main Number:
802-655-4040
Technical Assistance Center:
800-242-4685
Written Communication
If you prefer, you may write a letter with your comments and send it to:
®
Bio-Tek Instruments, Inc.
Technical Assistance Center
Highland Park, Box 998
Winooski, Vermont 05404-0998
USA
1-8
Introduction
Facsimile Support
You may send a fax with your questions or requests for help 24 hours a day to the following
numbers:
Technical Assistance Center:
802-655-3399
Electronic Communication
Electronic communication is available via the following:
E-Mail:
[email protected]
Internet Site:
www.biotek.com
PowerWaveX Operator's Manual
1-9
1-10
Introduction
Chapter 2
Installation
This chapter includes instructions for unpacking
and setting up the PowerWaveX, instructions for
connecting to printers and/or serial devices, and
descriptions of some of the reader’s components.
Operating Environment
For optimal operation, install the PowerWaveX on a level surface in an area where ambient
temperatures between 18°C (65°F) and 40°C (104°F) can be maintained. The reader is sensitive
to extreme environmental conditions. Conditions to avoid are:
•
Excessive humidity: Condensation directly on the sensitive electronic circuits can
cause the instrument to fail internal self-checks.
•
Excessive ambient light: Bright sunlight or strong incandescent light can reduce
the linear performance range and affect the instrument’s readings.
•
Dust: Optical density readings may be affected by extraneous particles (such as
dust) in the microplate wells. A clean work area is necessary to ensure accurate
readings.
Unpacking and Repackaging the Instrument
iii
Important! Keep the shipping cartons and the packaging material for the carrier’s
inspection. If the reader is shipped to the factory for repair or replacement, it must be
carefully repackaged using the original packing materials. Shipping with improper
packaging materials may void your warranty. If the original packaging materials
have been damaged, replacements are available from Bio-Tek.
1.
Carefully open the top of the shipping container, and remove the dust cover. See Figure 2-1.
The box will include a power cord, Operator’s Manual, serial cord, and optionally, a Service
Manual.
2.
Remove the top foam from the reader.
3.
Lift the reader out of the bottom foam, and place it on a level surface. Remove the reader
from the plastic bag.
4.
Place all shipping material back into the shipping box for reuse if the instrument needs to be
shipped again.
5.
Inspect the packaging and instrument for shipping damage such as visible dents or scratches.
6.
If the reader is damaged, notify the carrier and your manufacturer’s representative.
7.
Contact Bio-Tek's Technical Assistance Center for an RMA (Return Materials Authorization)
number before returning equipment for service. Mark the RMA number on the outside of the
shipping container. See page 1-8 for contact information.
Top foam
26 x32
2-Ml Poly Bag
Bottom foam
Figure 2-1: Unpacking and repackaging the reader
.
2-2
Installation
Removal of the Carrier Retention Screw in PowerWaveX Select Models
iii
Note: The PowerWaveX Select model is shipped with a carrier retention screw that
must be removed before the reader is used. An allen wrench (PN 49329) is supplied
for this task. See Figure 2-2 below.
Lift up the front door. Using the allen wrench provided, remove the carrier retention screw, wave
washer and flat washer. Save all of the shipping hardware.
iii
Important! Replace the carrier retention screw and washers prior to
shipment, or you will void the manufacturer’s warranty.
Carrier retention screw
(PN 19953)
Wave washer
(PN 17023)
Flat washer
(PN 17056)
Figure 2-2: Removal of the carrier retention screw and accompanying washers from the
PowerWaveX Select
PowerWaveX Operator's Manual
2-3
AFTER UNPACKING, VERIFY PERFORMANCE ____________________________________
Before using the PowerWaveX for the first time, verify that it is operating properly by turning it
on. Each time the reader is turned on, it performs a system self-test. If the self-test completes
successfully, the reader is ready for use. If the test fails, note any error codes and contact BioTek.
Refer to Chapter 4, Performance Verification/Qualification Tests for a recommended test
schedule, which includes Installation, Performance, and Operational Qualification Tests.
BEFORE REPACKAGING THE INSTRUMENT ______________________________________
1.
Decontaminate the reader prior to shipping (see Appendix A, Decontamination and Cleaning).
2.
For PowerWaveX Select models, replace the carrier retention screw and its accompanying
washers. See Figure 2-2 for an illustration.
3.
Once the reader is clean, pack it in its original shipping box, using original packing materials.
This shipping system was designed to be used no more than five times. If the container is
damaged and/or has been used more than five times, contact Bio-Tek for a new set of
shipping materials (PN 7263004).
Adjusting the Reader’s Wavelength Table Settings
The PowerWaveX has an internal wavelength table that stores six user-definable wavelength
values. When defining an assay, these six wavelengths are presented as selectable options. These
values can be easily viewed or changed:
•
From the Main Menu, select UTIL.
R E A D Y
R E A D
9 : 4 5 P M
D E F I N E
0 5 / 0 9 / 9
R E P O R T
U T I
9
L
The Main Menu
•
The Select Utility Option menu appears.
S E L E C T
T E S T S
U T I L I T Y
S E T U P
O P T I O N ?
O U T P U T
R E A D
Selecting SETUP from the Utility Option menu
2-4
Installation
•
From the Select Utility Option menu, select SETUP. The Edit Setup Information
menu appears.
E D I T
S E T U P
D A T E
T I M E
I N F O R M A T I O N ?
L A M B D A
* M O R
E
Selecting LAMBDA from the Edit Setup Information menu
•
From the Edit Setup Information menu, select LAMBDA.
•
The Enter Lambda #1 screen appears, showing the first value in the wavelength
table.
E N T E R
L A M B D A # 1
W A V E L E N G T H : 2 0 0
Viewing or changing the reader's internal wavelength table
•
To advance to the next wavelength, press ENTER.
•
To change the current wavelength value, use the reader's numeric keypad to enter a
number at the current cursor location. Once a selection is made, the cursor
automatically advances to the next editable location. To save the entry and advance
to the next wavelength, press the ENTER key.
•
Any wavelength in the range 200 to 999 nm may be entered (or 340 to 999 nm on
the PowerWaveX 340).
•
When the last (sixth) wavelength has been entered or viewed, the software returns to
the Edit Setup Information menu:
E D I T
S E T U P
D A T E
T I M E
I N F O R M A T I O N ?
L A M B D A
* M O R
E
Edit Setup Information menu
•
iii
Press the Main Menu key to return to the Main Menu.
Note: After selecting new wavelengths, the reader system test must be run.
From the Main Menu, press UTIL Î TESTS Î SYSTEM to start the test.
PowerWaveX Operator's Manual
2-5
Printing and Data Communications
INSTALLING A PRINTER ____________________________________________________
The parallel port on the PowerWaveX (see Figure 2-3 below) allows connection to Epson- and
HP-compatible deskjet printers. See Compatible Printers on page 2-17 for information on
specifying a printer. The reader’s parallel port requires a 25-pin D-sub-connector.
Parallel port
Monochromator
viewing port
Serial port
Figure 2-3: Serial and parallel connectors
The parallel port’s pinout definition is described in Table 2-1. A printer cable (PN 71072) is
supplied with the reader. If the cable that came with the reader gets lost or damaged, Bio-Tek
offers replacement printer cables.
Contact your authorized Bio-Tek dealer for information on cable prices and availability.
2-6
Installation
To attach a printer to the PowerWaveX:
1.
Power down the printer and place it in a location adjacent to the PowerWaveX.
2.
Attach one end of the parallel cable to the printer's parallel port.
3.
Attach the other end of the cable to the reader’s parallel port, located on the
instrument’s rear panel (see Figure 2-3).
4.
Ensure that the securing screws on both ends of the cable are tightened, and power
up the reader and printer.
iii
Important: To avoid system instability, connect the printer to the reader before
powering up the reader. Turn on the PowerWaveX first, then the printer.
PARALLEL PORT PIN DEFINITION _____________________________________________
Table 2-1 illustrates the pin definitions for the reader’s 25-pin (socket-female) D-sub Parallel
connector.
Table 2-1
Parallel Connector Pinouts
Parallel Port Pinout
Pin
1
2
3
4
5
6
7
8
9
10
11
12
13
PowerWaveX Operator's Manual
Signal
PSTROBE
D0
D1
D2
D3
D4
D5
D6
D7
NC
BUSY
NC
NC
Pin
14
15
16
17
18
19
20
21
22
23
24
25
Signal
NC
NC
RESET
NC
GND
GND
GND
GND
GND
GND
GND
GND
2-7
SERIAL PORT FOR COMMUNICATION WITH OTHER DEVICES ________________________
The PowerWaveX has a 25-pin serial (RS-232) port located on the rear panel of the instrument
(see Figure 2-3 for an illustration of the serial cable connection). The serial port allows the reader
to communicate with a computer, using standard communications software and/or RS-232
protocols.
Appendix B contains information on required protocols for computer control of the reader.
CONNECTING TO THE PC AND ESTABLISHING COMMUNICATION_____________________
1.
Power down the computer and the reader.
2.
Connect the appropriate serial cable to both machines (see Figure 2-3). The serial port on the
reader is a DTE configuration with a 25-pin (pin-male) D-sub connector. The connector’s
pinout is illustrated in Table 2-2.
3.
Power up the reader and the computer. Each time the reader is turned on, it performs a
system self test. If the self test completes successfully, the reader is ready for use. If the test
fails, note any error codes and contact Bio-Tek.
4.
Ensure that the PowerWaveX and the computer are operating with the same communications
parameters as described on the following page.
SERIAL PORT PINOUT DESCRIPTION ___________________________________________
Table 2-2 describes the reader’s serial /RS-232 pin connection.
Table 2-2
Serial Pinout Description
Serial Pin Description
Pin
1
2
3
4
5
6
7
8
9
10
11
12
13
2-8
Signal
NC
TX
RX
RTS
CTS
DSR
GND
DCD
NC
NC
NC
NC
NC
Pin
14
15
16
17
18
19
20
21
22
23
24
25
Signal
NC
NC
NC
NC
NC
NC
DTR
NC
RI
NC
NC
NC
Installation
SETTING UP COMMUNICATION PARAMETERS ____________________________________
Before serial communication can be initiated between the PowerWaveX and another device (such
as a host PC running Bio-Tek’s KC4 or KCjunior software packages), the communication
parameters must match on both devices. The reader’s default communication parameters are:
•
Baud Rate: 9600
•
Data Bits:
8
•
Parity:
None
•
Stop Bits:
2
The reader's Baud Rate can be changed to 1200 or 2400, if necessary. The reader's Data Bits,
Parity, and Stop Bits settings cannot be changed.
To change the Baud Rate:
•
At the Main Menu, select UTIL.
R E A D Y
9 : 4 5 P M
R E A D
D E F I N E
0 5 / 0 9 / 9
R E P O R T
U T I
9
L
Main Menu screen
•
At the Select Utility Option menu, select SETUP.
S E L E C T
T E S T S
U T I L I T Y
S E T U P
O P T I O N ?
O U T P U T
R E A D
Selecting Setup
•
At the Edit Setup Information screen, select *MORE to see more options, including
RS232.
E D I T
S E T U P
D A T E
E D I T
T I M E
S E T U P
R S 2 3 2
•
C A L
I N F O R M A T I O N
L A M B D A
* M O R
E
I N F O R M A T I O N ?
P L A T E
* M O R
E
Select RS232 to advance to the Select Baud Rate screen.
PowerWaveX Operator's Manual
2-9
•
The top line of the display shows the currently selected Baud Rate.
S E L E C T
B A U D
R A T E :
1 2 0 0
2 4 0 0
9 6 0 0
9 6 0 0
V I E W
Selecting the Baud Rate
•
To change the Baud Rate, press the soft key corresponding to the desired setting.
The top line of the display automatically updates to reflect the new setting.
•
To view the reader’s other communication settings, select VIEW.
R S 2 3 2
2
S E T T I N G S : N O
S T O P - B I T S
8
P A R I T
Y
D A T A - B I T
S
Viewing Other Communication Settings
2-10
Installation
Components
LAMP __________________________________________________________________
The lamp life is rated at an average of 1 billion flashes. This bulb should outlive the useful life of
the reader. If there is a problem with the lamp, however, the intensity may drop and the run-time
self check will detect a low signal level and generate an error message. If this happens, contact
Bio-Tek for bulb replacement information.
COOLING FAN ___________________________________________________________
Located on the bottom right edge of the instrument is a switch that activates an internal cooling
fan. When the PowerWaveX is on, the temperature in the measurement chamber will be
approximately 7°C above ambient temperature.
For users that need the measurement chamber to be close to ambient, Bio-Tek recommends using
the cooling fan. The fan will bring the temperature within 4°C of ambient. This would be
important (for example) for kinetic measurements that need to be made close to room temperature.
The fan should be off when non-temperature-sensitive testing is being completed, as prolonged
use may introduce dust into the internal mechanisms of the instrument.
PowerWaveX Operator's Manual
2-11
Adjusting the Line Voltage Input Range
The PowerWaveX is equipped with a four-voltage range power input module. This power input
module, located on the right side of the instrument, can be adjusted for 100 VAC, 115 VAC, 230
VAC and 240 VAC voltage inputs. The setting can be determined visually by observing which
indicator hole on the power input module has a peg within it. The ON/OFF switch and fuses are
also housed within the power input module. The following instructions can be used to change the
input voltage range. Figures 2-4A and B illustrate the power input module.
Figure 2-4A: Voltage selector board and fuse holder
Figure 2-4B: Selecting fuse position
1.
Unplug the reader and remove the power cord.
2.
Use a small flat-blade screwdriver to pop the fuse holder out of the power input
module.
2-12
Installation
3.
A small voltage select switchboard is located on the right of the power input
module. The switchboard, which can be removed with needle-nose pliers, can be
oriented four ways to select four different voltage input ranges.
4.
Once the switchboard is removed, text can be read on one side of the board. This
text identifies the switch positions. The text is always facing the ON/OFF switch
when installed in the power input module. The text matching the desired voltage
range should be on the edge of the switchboard, which goes into the power input
module as the arrow on the board indicates. The white plastic indicator will need to
rotate around so that it fits into the correct groove on the switchboard.
5.
Verify proper fusing before reinserting the switchboard and fuse holder.
6.
Once the switchboard is reinstalled, the white plastic indicator peg should line up
with the fuse holder voltage indicator holes. If the peg does not fit in the hole that
indicates the voltage intended, the switchboard is not installed correctly. Do not
power up the instrument until the voltage input range to be used is indicated
correctly by the peg.
PowerWaveX Operator's Manual
2-13
Adjusting the Fuse Configuration or Fuse Replacement
Both U.S. and European fuses are installed in the reader’s power input fuse module. The reader’s
fuses are configured at the factory prior to shipping. Use the following procedure if you need to
change the fuse configuration, or replace fuses. A failed fuse is usually an indication of another
problem, which a new fuse is not likely to fix. Contact Bio-Tek Technical Services if the fuse
replacement fails to rectify the problem.
1.
Use a small, flat-blade screwdriver to remove the fuse module from the power input
module.
2.
3.
The fuse module has two fuse configurations:
•
The U.S. configuration has a Hot fused .75-amp slo blo (PN 46023).
•
The European configuration has both Hot and Neutral fused
.315-amp slo blo T 5 X 20 mm (PN 46051).
To replace a defective fuse, pop out the old fuse and replace it with the correct new
one.
4.
The configuration of the fuse module (U.S. or European) is determined by which
fusing network is facing the inside of the power input module (refer to Figure
2-4B). To change the configuration, remove the Phillips-head screw that anchors the
fuse holder. Remove the fuse holder and turn it over. Replace the screw and
reinstall the fuse module in the power input module.
2-14
Installation
Other Utility Options
The PowerWaveX contains several global configurable options, such as date and time, report
output, and plate reading preferences. These options are accessed via the Select Utility Option
menu (see below), and include:
•
TESTS: Run a SYSTEM test to check reader optics or the CALPLATE test to check
alignment and calibration. Select CHKSUM to perform an internal checksum test,
and to display the version and configuration information for the currently installed
software.
•
SETUP: Set the current date and time, as well as the date and time formats.
•
OUTPUT: Specify where plate data should be sent, to a printer, a computer, or to
both. Additional options include report format (Column and/or Matrix), and whether
or not to print standard curves.
•
READ: Enable or disable read-time prompting for Plate ID, Sample ID, and Sample
Count. Specify whether or not to read in Rapid mode.
(MAIN)
READ
DEFINE
REPORT
UTIL
↓
SELECT UTILITY OPTION:
TESTS
SETUP
OUTPUT
READ



↓



PROMPT FOR PLATE ID?: YES



YES


↓


REPORT OUTPUT: PRINT


PRINT

↓
COMPUTER

EDIT SETUP INFORMATION:

DATE
TIME
NO
LAMBDA
BOTH
*MORE
↓
SELECT TEST:
SYSTEM
CHKSUM
CALPLATE
Figure 2-5: Options available under UTIL
PowerWaveX Operator's Manual
2-15
SETTING THE DATE AND TIME _______________________________________________
To set the current Date and Time, and/or to change their formats:
•
From the Main Menu, select UTIL Î SETUP.
The Edit Setup Information menu will appear.
E D I T
S E T U P
D A T E
I N F O R M A T I O N
T I M E
L A M B D A
* M O R
E
M D
Y
Selecting DATE from the Edit Setup Information menu
•
Select DATE. The DATE entry screen will appear.
D A T E :
0 3 / 1 6 / 9 9
M M D D Y Y
D D M M Y Y
Entering the Date and selecting a format
•
Enter the date using the numeric keys on the keypad. The cursor is positioned under
the first editable field, and advances automatically.
•
To change the date format, select MMDDYY or DDMMYY. The display automatically
updates to reflect the new format.
•
Press ENTER to return to the Edit Setup Information menu.
•
To change the current time and/or the time format, select TIME from the
Edit Setup Information menu. The Time entry screen will appear.
T I M E :
1 2 H O U R
0 3 : 1 1 P M
2 4 H O U R
1 2 H
R
A M / P
M
Entering the Time and selecting a format
•
Enter the time using the numeric keys on the keypad. The cursor is positioned under
the first editable field, and advances automatically.
•
To change the time format, select 12HOUR or 24HOUR, then AM or PM. The
display automatically updates to reflect the new format.
•
2-16
Press ENTER to return to the Select Utility Options menu.
Installation
SPECIFYING DATA OUTPUT AND REPORTING OPTIONS_____________________________
To specify data output options:
•
From the Main Menu, select UTIL Î SETUP Î OUTPUT. The Report Output
screen will appear.
Data Output
R E P O R T
P R I N T
O U T P U T ?
P R I N T
C O M P U T E R
B O T H
Selecting the Report Output
•
The current output option is displayed on the top line. Select PRINT to send reports
directly to a printer, COMPUTER to send data out through the serial port, or BOTH.
•
Note: These options have no effect on data output if the reader is being controlled
by software (such as KC4 or KCjunior) running on a host PC.
•
Press ENTER to continue. The Select Printer screen will appear.
Compatible printers
S E L E C T
E P S O N
P R I N T E R :
E P S O N
H P
Selecting a printer
•
BTI readers support printers using either HP's PCL3 language, such as the HP
DeskJet series, or Epson's LQ language. For the latest list of compatible printers,
contact Bio-Tek Instruments' Technical Assistance Center (see page 1-8 for details),
or visit our website, www.biotek.com.
•
Select EPSON or HP as appropriate.
•
Press ENTER to continue. The Report Type screen will appear.
PowerWaveX Operator's Manual
2-17
Report Types
R E P O R T
T Y P E ?
C O L U M N
M A T R I X
M A T R I X
B O T
H
Selecting the Report Type
•
Note: Appendix D contains sample reports.
•
The currently selected report type is displayed in the top line. Select COLUMN to
print information in a list (columnar) format, MATRIX to print in a format that
resembles the plate type (e.g., 8 x 12 matrix), or BOTH.
•
Press ENTER to continue. The Samples in Col Rpt screen will appear.
S A M P L E S
Y E S
I N
C O L
R P T ?
N O
N O
Specifying whether or not to include samples on the Column Report
•
Select YES to print results for all wells on the plate, including samples.
•
Select NO to limit the results information to blanks, controls, and standards.
•
Press ENTER to continue. The Print Curve-Fit screen will appear.
P R I N T
C U R V E - F I T ?
Y E S
N O
N O
Specifying whether or not to print the standard curve
•
Select YES to print the standard curve (if the assay generates one), or NO to suppress
this report.
•
2-18
Press ENTER to return to the Select Utility Option menu.
Installation
SELECTING READ OPTIONS _________________________________________________
To specify various read-time options:
•
From the Main Menu, select UTIL Î READ.
The Prompt for Plate ID screen will appear. Press ENTER to cycle through the
prompt screens.
P R O M P T
F O R
Y E S
I D ?
Y E
S
N
O
Y E
S
N O
P R O M P T
F O R
Y E S
S A M P L E
I D ?
N O
P R O M P T
S A M P L E
Y E S
R E A D
P L A T E
C O U N T ?
N O
I N
Y E S
R A P I D
M O D E ?
N O
N O
Selecting Read Preferences
•
If selected, at read-time:
PLATE ID prompts for microplate identification.
SAMPLE ID prompts for an identification for each sample group.
SAMPLE COUNT prompts for the number of samples on the plate.
•
If RAPID MODE is disabled, the plate reads at a speed designed to ensure low CVs
and precise results. If Rapid Mode is enabled, the plate reads more quickly but may
possibly result in higher CVs. If an assay favors speed over precision, consider
reading in Rapid Mode.
•
Pressing ENTER after each selection advances the display.
•
When selections are completed, the display returns to the Select Utility Option
menu.
PowerWaveX Operator's Manual
2-19
2-20
Installation
Chapter 3
Operation
This chapter includes instructions for operating
the PowerWaveX and its software.
Overview
The PowerWaveX features a 25-key front panel with a 2 X 24 character LCD (Liquid Crystal
Display), allowing access to the reader’s program menus. Test results can be sent to an attached
printer and/or through the reader’s bi-directional serial port to a host computer. The serial port also
allows for computer control of the instrument, and provides the means for downloading additional
assay definition files to the instrument.
The PowerWaveX may be loaded with custom pre-programmed assays and/or with “Open” assays.
•
The pre-programmed assays allow you to easily select, modify, and run
(for example), several common ELISA and important nucleic acid applications
directly from the front panel.
•
The Open assays contain standard default settings, upon which assays can
be programmed.
This chapter describes how to program Open assays on the PowerWaveX. The custom
pre-programmed assays are described in Bio-Tek's Assay Reference Guide (PN 7271006).
iii
Important: Screens and options throughout this chapter pertain to “Open” assays.
Pre-programmed assays may not show all possible screens or options, or may show
custom screens not described here. Please refer to the Assay Reference Guide for
details on pre-programmed assays.
Front Panel
LCD
READY
READ
10:23AM
37.0°C
DEFINE REPORT UTIL
Menu functions
Softkeys (4).
Press a key to
choose a menu
function.
Press the left and right
arrows to move the
cursor in the display.
Press Options to
scroll through options
or the alphabet.
Press Shift + Options
to scroll backward
through options.
Figure 3-1: PowerWaveX Front Panel
SELECTING MENU FUNCTIONS _______________________________________________
The menu functions appear in the bottom (second) line of the front panel's LCD. To make a
selection, press the SoftKey below the menu function.
SCROLLING THROUGH OPTIONS _______________________________________________
Certain functions, such as entering an assay name or selecting an assay, offer a set of options from
which to make a selection. To view the different options, press the Options key or the Shift +
Options key combination. Press the ENTER key to select the current option.
3-2
Operation
ENTERING ASSAY NAMES ___________________________________________________
NAME:
-
DNA Quant
/
:
_
Each PowerWaveX assay requires the entry of a name, using up to 16 alphanumeric characters. To
enter an assay name via the keypad, press Shift + key A-H, or scroll through the alphabet with the
Options key for A-Z. Use the forward or reverse arrows to move the cursor within the display. To
delete an entry, press the CLEAR key. Press the ENTER key to store the completed assay name and
continue.
The NAME display offers four symbols that can be used in an assay name: dash (hyphen), forward
slash, colon, and underscore. These symbols appear in the LCD as SoftKey choices (see above).
To include a symbol within an assay name, press its corresponding SoftKey.
MOVING BACKWARD THROUGH THE MENUS ____________________________________
To move to a previous menu, press the Previous Screen key.
SAVING ASSAYS __________________________________________________________
After defining an assay, press the Main Menu key. The changes are saved automatically.
STARTING OR STOPPING A READ ______________________________________________
To begin a plate read, press the READ key. To abort the read, press the STOP key.
Startup Screen
The PowerWaveX performs a self-test when powered on, displaying the Startup screen until
initialization is complete. During this period, no keys are active. If the instrument fails to pass the
self-test, a beep will sound, and an error code will display. Refer to Appendix C to interpret error
codes. Contact Bio-Tek Instrument’s Technical Assistance Center for assistance on troubleshooting
errors (see page 1-8 for contact information).
I N S T R U M E N T
I D
S E L F - T E S T . . .
PowerWaveX startup screen
PowerWaveX Operator’s Manual
3-3
Main Menu
Once the system is initialized, the Main Menu is displayed. The keypad’s four SoftKeys, located
below the on-screen menu options (READ, DEFINE, REPORT and UTIL), are activated, and may
be selected.
R E A D Y
9 : 4 5 A M
R E A D
D E F I N E
0 5 / 0 9 / 9
R E P O R T
U T I
9
L
Main Menu screen
Press the SoftKey that corresponds to a displayed menu option to activate that option:
•
READ: Choose an assay for plate reading. Alternatively, press the key labeled
READ on the keypad.
•
DEFINE: Define the data reduction parameters for a new assay, or modify an
existing one.
•
REPORT: Print a Result, Map, Assay, or List report.
•
UTIL: Run a System Test or Calibration Plate Test. Set up various global
configuration options such as date and time, report output, and plate reading
preferences.
(MAIN)
READ
DEFINE
REPORT
UTIL






SELECT UTILITY OPTION:



TESTS


↓


PRINT REPORT:


RESULT

↓
↓
MAP

SELECT ASSAY NUMBER: 01

NAME: RAW_OD
SETUP
OUTPUT
ASSAY
LIST
READ
↓
SELECT ASSAY NUMBER: 01
NAME: RAW_OD
Figure 3-2: Options available from the Main Menu
3-4
Operation
DEFINE
The Main Menu option DEFINE allows you to define the data reduction parameters for a new
assay, or to modify a previously defined assay stored in memory.
(MAIN)
READ
DEFINE
REPORT
UTIL
↓
SELECT ASSAY NUMBER: 01
NAME: RAW_OD
Press Options to scroll through the assay list.
Press ENTER to select an assay.
↓
NAME: RAW_OD
-
/
:
SPACE
See Editing the Assay Name on page 3-7 for instructions on modifying the current
name.
Press ENTER to continue.
↓
DEFINE:
METHOD
MAP
FORMULA
CURVE
Figure 3-3: Options available under DEFINE
•
METHOD: Define the method of reading, such as Endpoint, Kinetic, or Scan, and
Single or Dual Wavelength. Specify a delay before read, or incubation settings.
•
MAP: Specify the plate layout, using blanks, controls, standards, and/or samples.
Choose to map the plate manually, or let the software map it automatically.
•
FORMULA: Define cutoff, transformation, validation, and/or general formulas. Create
variables to be used within formulas.
•
CURVE: Specify a curve fit type and x/y axis types (lin/log). Specify the method by
which standard outliers can be edited. Enable or disable the extrapolation feature.
PowerWaveX Operator’s Manual
3-5
SELECTING AN ASSAY ______________________________________________________
From the Main Menu, press the soft key beneath the DEFINE menu option to access the Select
Assay Number screen.
S E L E C T
A S S A Y
N U M B E R : 0 1
N A M E : H B S -A G 1
Assay Selection screen showing the current assay name and number
•
Use the NUMERIC keys to enter the number of any predefined assay stored in the
reader’s memory, or the OPTIONS key to advance one assay at a time. The cursor is
positioned at the first editable field, and advances automatically. The numeric range
depends on the number of assays programmed in the reader’s memory. (The reader
typically has 55 “open” assays available.)
•
Press ENTER to advance to the Edit Assay Name screen. You may change the default
assay number to a more descriptive one (see Editing the Assay Name on the following
page):
CLEAR: Clears the reader’s display.
MAIN MENU: Returns the display to the Main Menu screen.
PREVIOUS SCREEN: Returns the display to the previous screen.
ENTER: Saves the current settings and advances to the next screen.
iii
3-6
Note: Within every screen, these keys will continue to function as
described above.
Operation
EDITING THE ASSAY NAME __________________________________________________
Use the Edit Name screen to edit the name currently assigned to the assay. The assay name can
contain up to 16 alphanumeric characters.
N A M E :
H B S - A G 1
-
/
:
S P A C
E
Edit Assay Name screen
•
The cursor is positioned at the first editable field (e.g., under “H”).
•
Use the ALPHA and NUMERIC keys to change the Assay name.
•
Use the OPTIONS key to sequentially advance the character positioned above the
cursor. The characters will cycle through the alphabet (A-Z), with a space following
Z.
•
Use the LEFT◄ and RIGHT►arrow keys to move the cursor to the previous or next
editable field. The cursor will wrap around the edit field.
•
Use SOFT KEYS 1, 2, 3 and 4 to select a dash, forward slash, colon, or space for
inclusion in the assay name.
PowerWaveX Operator’s Manual
3-7
Define METHOD
To define read method parameters, start at the Main Menu and select DEFINE, select the assay,
press ENTER, then select METHOD.
D E F I N E
M E T H O D
M A P
F O R M U L A
C U R V
E
Define options screen
The definable read method parameters include:
iii
•
Endpoint, Kinetic or Well Scanning Read Modes
•
Delay first read
•
Incubation parameters
•
Wavelengths applied
•
Shake parameters
•
Kinetic analysis
•
Number of points for well scanning
•
Spectral scan (Assay 00 only)
Note: Some screens described in this section may not appear on certain reader
models.
READ TYPE ______________________________________________________________
Use the READ TYPE screen to specify the reading method, Endpoint, Kinetic, or Scan.
R E A D
T Y P E : K I N E T I C
E N D P O I N T
K I N E T I C
S C A N
Read Type screen
•
During an ENDPOINT read, the plate is read once with one (single) or two (dual)
wavelengths. One optical density (OD) value is reported for each well.
3-8
Operation
•
During a KINETIC read, the plate is read a defined number of times with one (single)
or two (dual) wavelengths. Three different types of data are available (see Kinetic
Analysis on page 3-15):
For a rate determination, the data calculated is the rate of OD change. After
determining the maximum rate of change per well using linear regression, the
units are milliOD/min.
The R-squared value is an indication of how well the points fit the linear
curve.
The Onset time indicates how many seconds it took to reach a user’s
predetermined OD value.
•
During WELL SCANNING, the Optical Density values are measured at different
points across the well diameter. Choose from 1 to 31 points total (15 left of center, 15
right of center). If you choose 1, then the plate is read as an endpoint assay. The data
out is equal to the sum of all of the OD values recorded for the well. To find the
average OD value for the region scanned, divide the scanned OD result by the total
number of scans. For more information, see Well Scanning on page 3-16.
Spectral Scanning
Well Scanning and Spectral Scanning are two different read methods.
Certain readers contain the pre-programmed assay SPECTRAL SCAN (number 00). This assay
allows for the definition of start, stop, and step (increment) wavelength values. For example, if the
Start/Stop/Step values are 200/600/10 nm, respectively, each well will be read at 200, 210, 220 …
600 nm.
The maximum number of wavelengths that can be specified for reading within an assay is 100. The
printed report shows up to 100 points and includes the Peak OD and wavelength at Peak OD.
PowerWaveX Operator’s Manual
3-9
DELAY FIRST READ ________________________________________________________
The Delay First Read option can be used to allow for sufficient time for microwell contents to settle,
or to incubate a plate before reading.
D E L A Y
F I R S T
R E A D
T I M E :
M M : S S
Delay in first read entry
•
Use the NUMERIC keys to enter the time in minutes and seconds. The maximum
delay time is 59 minutes, 59 seconds (59:59).
INCUBATION TEMPERATURE (READER-DEPENDENT)_______________________________
Certain reader models are equipped with heaters to perform plate reads at elevated temperatures (see
Chapter 1, Introduction for information on these models). The Incubation Temperature screen is
used to enable incubation for this assay, and to set the temperature.
I N C U B A T I O N
A M B I E N T
T E M P :
3 7 ° C
T E M P E R A T U R E
Incubation Temperature entry
•
Select AMBIENT to read the plate at room temperature.
Note: See Cooling Fan in Chapter 2 for information on lowering the ambient
temperature by turning on the internal fan.
•
Select TEMPERATURE to read the plate at an elevated temperature.
•
If TEMPERATURE is selected, use the NUMERIC keys to enter the incubation
temperature. The valid range is from 25 to 50° C. Incubation performance is
specified from 4° over ambient to 50° C.
3-10
Operation
WAVELENGTH(S)__________________________________________________________
Use the Wavelength selection screen to specify whether the plate will be read at one (SINGLE) or
two (DUAL) wavelengths.
W A V E L E N G T H :
S I N G L E
D U A L
D U A L
Specifying the Wavelength mode, Single or Dual
•
If SINGLE is chosen, the reader measures the optical density of each well with a single
wavelength.
•
If DUAL is chosen, each well is read twice, at two different wavelengths. The
microplate is not removed from the reading chamber between the two measurements.
The final reported optical density is the difference between the two readings.
iii
Note: Dual wavelength readings can significantly reduce optical interference caused
by scratches or fingerprints that absorb light equally at both wavelengths. Choose a
REF wavelength that is far from the MEAS wavelength in the spectrum for best
results.
MEASUREMENT WAVELENGTH(S) _____________________________________________
After specifying Single or Dual Wavelength, the MEAS or MEAS/REF selection screen appears,
for wavelength selection.
M E A S : 4 0 5
4 0 5
4 5 0
M E A S : 4 5 0
4 0 5
4 9 0
* M O R E
R E F : 6 3 0
4 5 0
4 9 0
* M O R E
Specifying the wavelength(s) to be used during plate reading. Single wavelength selection is
shown on top, Dual wavelength selection on bottom.
PowerWaveX Operator’s Manual
3-11
•
If reading at a single wavelength, a MEASurement wavelength must be selected. If
reading at dual wavelengths, a REFerence wavelength must also be selected.
•
Choose a wavelength value from those presented. Select *MORE to see additional
wavelengths.
•
Note: If a desired wavelength is not listed, the reader's internal wavelength table must
be changed. Press the Main Menu button to save the current assay settings and return
to the Main Menu. See Adjusting the Reader's Wavelength Table Settings in Chapter
2 for instructions.
KINETIC READ PARAMETERS _________________________________________________
If the Kinetic read type was specified, a few accompanying parameters must be defined, including
kinetic method, reading interval, and number of reads.
K I N E T I C :
T O T A L
T O T A L
R E A D S
R E A D S
D U R A T I O N
Kinetic reading method selection screen
•
Select TOTAL READS to read the plate a specified number of times.
•
Select DURATION to read the plate for a specified length of time.
Kinetic Interval
Regardless of the kinetic reading method selected (Total Reads or Duration), a Kinetic Interval is
required. This is the length of time (in hours, minutes and seconds), between each kinetic read.
K I N E T I C
I N T E R V A L :
H H : M M : S S
Kinetic Interval entry screen
•
Use the NUMERIC keys to enter the interval.
•
The valid entry ranges are: 0-1 hours, 0-59 minutes, and 0-59 seconds.
•
Note: If the kinetic reading method is DURATION, the Interval/Duration combination
must result in 2 to 40 total kinetic reads.
3-12
Operation
Total Number of Kinetic Reads
If the TOTAL READS kinetic reading method was selected, specify the total number of kinetic reads.
K I N E T I C
N U M B E R
T O T A L
O F
R E A D S :
1 0
Kinetic Reads entry screen
•
Use the NUMERIC keys to enter the total number of reads to perform.
•
The valid range is 2 to 40 reads.
Kinetic Duration
If the DURATION kinetic reading method was selected, specify the total length of time during which
readings will be taken.
K I N E T I C
D U R A T I O N :
H H : M M : S S
Kinetic Duration entry screen
•
Use the NUMERIC keys to enter the time duration in hours, minutes and seconds.
•
The maximum duration time is 80 hours. Note: The Kinetic Interval/ Duration
combination must result in 2 to 40 total kinetic reads.
PLATE SHAKING __________________________________________________________
A variety of plate shaking options are available on the PowerWaveX reader.
S H A K E : B E F O R E
E V E R Y
F I R S T
N O N E
E V E R Y
R E A D
Shake Mode Selection screen
•
For non-kinetic reads, choose FIRST to enable shaking, or NONE to indicate no
shaking.
•
For kinetic reads, choose FIRST to shake before the first read only, EVERY to shake
before every read, or NONE to indicate no shaking.
PowerWaveX Operator’s Manual
3-13
Shake Time
If shaking is enabled, a Shake Time must be specified.
S H A K E
T I M E :
H H : M M : S S
C O N T I N U O U S
Shake Time entry screen
•
Use the NUMERIC keys to enter the shake duration. Valid ranges are: 0-1 hours, 0-59
minutes, and 0-59 seconds.
•
For a kinetic read with Shake Before Every Read enabled, the CONTINUOUS option
is available. If selected, the shake duration is set automatically, according to the
kinetic interval.
Shake Speed
For the PowerWaveX, the shake movement is a repeated 0.021-inch movement from the shake
position and back. Choose from several different speeds, including a Variable option.
S H A K E
L O W
S P E E D :
M E D I U M
M E D I U M
H I G H
V A R I
Shake Speed selection screen
3-14
•
Select LOW for low-speed (17 Hz) shaking.
•
Select MEDIUM for medium-speed (18 Hz) shaking.
•
Select HIGH for high-speed (19 Hz) shaking.
•
Select VARI variable-speed shaking (1 second of each speed repeated).
Operation
KINETIC ANALYSIS ________________________________________________________
For kinetic reads there are three calculation options to choose from.
K I N E T I C
R A T E
A N A L Y S I S :
R - S Q R
R - S Q R
O N S E T
Kinetic Data Analysis selection screen
•
Select RATE to apply a linear fit to calculate the maximum slope in mOD/min. based
on the number of kinetic points specified.
•
Select R-SQR to calculate the R-squared value at the maximum slope, based on the
linear curve fit and the number of kinetic points specified.
•
Select ONSET to calculate the time it takes for each well to reach the specified onset
optical density.
Kinetic Points
For the kinetic analysis selections RATE and R-SQR, specify the number of sequential kinetic
points used to calculate the steepest Rate or the R-squared value at the steepest Rate.
K I N E T I C
A L L
P O I N T S :
0 3
P O I N T S
Defining the number of Kinetic Points for use in calculations
•
Select ALL to indicate all of the kinetic reads.
•
To specify a subset of kinetic reads, select POINTS, then use the NUMERIC keys to
input the number of points. The valid entry range is 2 to the total number of kinetic
reads.
iii
Note: If the number of kinetic reads is changed during assay definition, you may need
to adjust the Kinetic Points setting. The number of Kinetic Points must be less than or
equal to the total number of kinetic reads.
PowerWaveX Operator’s Manual
3-15
Onset OD
For the Kinetic Analysis selection ONSET, an Onset OD is required.
E N T E R
O N S E T
O D :
2 . 5 0
0
Entering the Onset OD for kinetics
•
Use NUMERIC keys to enter the onset OD. The valid entry range is 0.000 to 3.000
OD.
WELL SCANNING __________________________________________________________
For the Scanning Read Type, multiple readings are taken across each microwell. The number of
readings, or Scan Points, is configurable.
E N T E R
O F
N U M B E R
S C A N
P O I N T S ?
1 1
Entering the number of scan points per well
•
The maximum number of Scan Points is 31. Only odd integers are accepted, to ensure
that the center of the well is always read.
•
The 31 scan positions are fixed in the software. You must determine the optimal
number of scans per well. If, for example, 7 scans across the well is chosen, the reader
will read the centermost seven points in the well. The more scan points chosen, the
closer to the well sides reads will be taken.
•
iii
The reader reports the sum of OD values for all points scanned.
Note: Exercise caution when specifying the Scan Points value. If too many points are
specified, the sides of the wells may be getting read, possibly resulting in an
artificially high reading.
3-16
Operation
Define MAP
To configure the plate layout, start at the Main Menu and select DEFINE, select an assay, press
ENTER, then press MAP.
D E F I N E
M E T H O D
M A P
F O R M U L A
C U R V
E
Selecting the Map option on the DEFINE screen
The following MAP parameters can be defined/modified:
•
Automatic or Manual Map Generation
•
Mapping Direction
•
Replication Direction
•
Blank Map Selection
•
Blanking Constant
•
Number of Blanks
•
Location of Blanks
•
Number of Standards
•
Number of Standard Replicates
•
Averaging of Standards
•
Concentration and Location of Standards
•
Number of Controls
•
Control Type Definition
•
Number of Control Replicates
•
Control Location
•
Number of Samples
•
Number of Sample Replicates
•
Sample Location
PowerWaveX Operator’s Manual
3-17
MAP GENERATION _________________________________________________________
“Map Generation” represents the method by which blanks, controls, standards, and/or samples are
assigned to specific locations on the plate.
M A P
G E N E R A T I O N :
A U T O
M A N U A L
M A N U A L
Selecting Manual or Automatic map generation
•
Select AUTO to instruct the software to automatically generate a Plate Map after the
blanks, controls, standards and/or samples have been defined.
•
Select MANUAL to indicate that the well assignments will be performed manually (by
the user) at Define and/or Run time.
iii
Note: Press Shift + Clear at the MAP GENERATION screen to “clear” a previously
defined map.
Mapping Direction
The “Mapping Direction” is the direction (Down or Across) in which blank, control, standard, or
sample groups will be mapped on the plate.
M A P P I N G
D O W N
D I R E C T I O N : D O W N
A C R O S S
Selecting the direction for group mapping
•
Select DOWN to map groups down the plate columns.
•
Select ACROSS to map groups across the plate rows.
•
For example, if the plate contains one blank (BLK), one negative control (NC), one
positive control (PC), and samples with one replicate:
DOWN maps the wells as A1:BLK, B1:NC, C1:PC, D1:SMP1, E1:SMP2, and
so on.
ACROSS maps the wells as A1:BLK, A2:NC: A3:PC, A4:SMP1, A5:SMP2,
and so on.
•
3-18
Additional examples of mapping directions are shown in Figure 3-4.
Operation
Replication Direction
The “Replicate Direction” is the direction (Down or Across) in which blank, control, standard, or
sample replicates will be mapped on the plate.
R E P
D O W N
D I R E C T I O N : A C R O S S
A C R O S S
Selecting the direction for replicate mapping
•
Select DOWN to map replicates down the plate columns.
•
Select ACROSS to map replicates across the plate rows.
•
For example, if the plate contains two blanks (BLK), two negative controls (NC), two
positive controls (PC), and samples in duplicate:
DOWN maps the wells as A1/B1:BLK, C1/D1:NC, E1/F1:PC, G1/H1:SMP1.
ACROSS maps the wells as A1/A2:BLK, A3/A4:NC, A5/A6:PC,
A7/A8:SMP1, and so on.
•
Additional examples of mapping directions are shown on page 3-21.
PowerWaveX Operator’s Manual
3-19
Map Direction DOWN, Rep Direction DOWN:
A
B
C
D
E
F
G
H
1
STD1
STD1
STD2
STD2
STD3
STD3
STD4
STD4
2
STD5
STD5
PC
PC
NC
NC
SMP1
SMP1
3
SMP2
SMP2
SMP3
SMP3
SMP4
SMP4
SMP5
SMP5
4
5
6
7
8
9
10
11
12
7
STD4
SMP3
8
STD4
SMP3
9
STD5
SMP4
10
STD5
SMP4
11
PC
SMP5
12
PC
SMP5
7
8
9
10
11
12
7
NC
NC
8
SMP1
SMP1
9
SMP2
SMP2
10
SMP3
SMP3
11
SMP4
SMP4
12
SMP5
SMP5
Map Direction ACROSS, Rep Direction ACROSS:
1
A STD1
B NC
C
D
E
F
G
H
2
STD1
NC
3
STD2
SMP1
4
STD2
SMP1
5
STD3
SMP2
6
STD3
SMP2
Map Direction DOWN, Rep Direction ACROSS:
A
B
C
D
E
F
G
H
1
STD1
STD2
STD3
STD4
STD5
PC
NC
SMP1
2
STD1
STD2
STD3
STD4
STD5
PC
NC
SMP1
3
SMP2
SMP3
SMP4
SMP5
4
SMP2
SMP3
SMP4
SMP5
5
6
Map Direction ACROSS, Rep Direction DOWN:
1
A STD1
B STD1
C
D
E
F
G
H
2
STD2
STD2
3
STD3
STD3
4
STD4
STD4
5
STD5
STD5
6
PC
PC
Figure 3-4: Examples of Mapping Directions
3-20
Operation
Start Mapping at Well Location
Enter the location of the well that will be the starting point for automatic mapping.
S T A R T
A T
M A P P I N G
W E L L
L O C A T I O N :
A 0 1
Entering the starting well for automatic plate mapping
•
Use the NUMERIC and ALPHA keys to enter a letter or number at the cursor location.
The valid entry range is from A01 to H12, depending on the number of blanks,
standards, controls, and/or samples defined in the assay.
Blank Map
There are several blanking methods to choose from.
B L A N K
A I R
M A P :
F U L L
B L A N K
R O W
M A P :
F U L L
C O N S T
F U L L
C O L U M N
B L A N K
M A P :
P - A C R O S S
* M O R E
* M O R E
F U L L
P - D O W N
* M O R E
Selecting a blanking method
•
AIR performs an initial reading “on air” just prior to the plate read, and uses that
reading as the blank value. This value is subtracted from each well on the plate.
•
FULL enables a single blank well or an average of blank wells (up to 48) to be
subtracted from each well on the plate.
•
CONST (Constant) allows entry of a user-specified absorbance value. This value will
be subtracted from each well on the plate.
PowerWaveX Operator’s Manual
3-21
•
ROW enables a single blank well or an average of blank wells to be selected for each
row. The maximum number of blanks is 48. Use manual mapping to position blanks,
standards, controls, and samples.
•
COLUMN enables a single blank well or an average of blank wells to be selected for
each column. The blank OD or average OD will be subtracted from other wells in the
column. Use manual mapping to position blanks, standards, controls, and samples.
•
P-ACROSS enables a blank in every even-numbered column to be subtracted from the
well to the left of it in every odd column. Use manual mapping to set up the
appropriate map by placing the standards, controls, and samples in only the odd
columns.
•
P-DOWN enables a blank in the B, D, F and H rows to be subtracted from the well
above in the A, C, E and G rows. Use manual mapping to set up the appropriate map
by placing the standards, controls, and samples in rows A, C, E, and G.
Blanking Constant
If the CONST blanking method is selected, a Blanking Constant must be defined. This value will be
subtracted from each well on the plate.
E N T E R
B L A N K I N G
C O N S T A N T :
0 . 5 0 0
Entering the Blanking Constant value
•
Use the NUMERIC keys to enter the value. The valid entry range is 0.000 to 3.000
OD.
Number of Blanks
If the FULL, COLUMN, or ROW blanking method is selected, enter the number of blank wells on the
plate.
E N T E R
N U M B E R
O F
B L A N K S :
0 2
Entering the number of blank wells on the plate
•
Use the NUMERIC keys to enter the number of blank wells. The valid entry range is 0
to 48 wells.
3-22
Operation
Blank Location
The Blank Location screen is presented if blank wells are defined and manual map generation is
selected.
E N T E R
T H E
L O C A T I O N
O F
B L A N K # 1 :
A 0 1
Defining the Location of the blank wells
•
Use the NUMERIC and ALPHA keys to enter the location of the first Blank well.
•
Press ENTER to define subsequent Blank well location(s).
Number of Standards
For assays requiring standards, begin by entering the number of standard groups.
E N T E R
N U M B E R
O F
S T A N D A R D S :
0 5
Entering the number of standard groups
•
Use the NUMERIC keys to enter the number of standard groups.
Enter 00 to indicate no standards.
•
The valid entry range depends on the selected curve fit method. The maximum number
of standards for all curve fit methods is 12. The minimum numbers of standards are:
4 for 2-P, 4-P, cubic, cubic spline
3 for quadratic
2 for linear, point-to-point
•
Note: If the NUMBER OF STANDARDS setting is modified, the number of standard
replicates (see next page) automatically defaults to 01.
PowerWaveX Operator’s Manual
3-23
Number of Standard Replicates
The Standard Replicates entry screen is presented if the number of standard groups is greater than 0.
E N T E R
N U M B E R
S T A N D A R D
O F
R E P L I C A T E S :
0 2
Entering the number of standard replicates
•
Use the NUMERIC keys to enter the number of standard replicates. The valid entry
range is from 1 to 8 replicates.
The software will automatically verify that the number of replicates,
multiplied by the number of standards, does not exceed the number of wells
on the plate.
Average Standards
If the Number of Standard Replicates setting is greater than 01, the Average Standards? screen is
presented.
A V E R A G E
Y E S
S T A N D A R D S ?
Y E S
N O
Choosing whether or not to Average Standards
•
Select YES to average the replicates for each standard group, and then use the group
averages when calculating the standard curve.
•
3-24
Select NO to use the individual standard replicates when calculating the standard curve.
Operation
Standard Concentrations
Use the Standard Concentration screen to enter the predicted concentration value for each standard
group. If manual map generation is selected, the replicate locations must also be defined.
C O N C .
O F
L O C A T I O N
S T D # 1 : 1 . 5 0
R E P # 1 : A 0
1
Entering the predicted standard concentrations, and well locations (if manual map)
•
Use the NUMERIC, ALPHA, and DECIMAL POINT keys to enter the standard
concentration values. The valid entry range is .00001 to 999999. The entry cannot
exceed 6 characters, including the decimal point.
•
If automatic mapping is selected, each replicate's location is available for viewing
only. Pressing ENTER advances to the concentration value entry for the next standard.
•
If manual mapping is selected, the location must be defined. Pressing ENTER from
the standard concentration entry moves the cursor to the Location field. Pressing
ENTER from the Location field advances to the concentration value entry for the next
standard.
Number of Controls
For assays requiring controls, begin by entering the number of control groups.
E N T E R
N U M B E R
O F
C O N T R O L S :
0 2
Entering the number of control groups
•
Use the NUMERIC keys to enter the Number of Control groups in the assay. For
example, if the assay requires one or more positive control wells and one or more
negative control wells, enter 02.
•
The valid entry range depends on the number of locations on the plate that are
undefined. The maximum number of control groups is 8.
PowerWaveX Operator’s Manual
3-25
Control Type
After defining the number of controls for this assay, select the types of controls to use.
C O N T R O L # 1 : P C
P C
N C
H P C
* M O R
E
* M O R
E
* M O R
E
C O N T R O L # 1 : P C
L P C
C T L 1
C T L 2
C O N T R O L # 1 : P C
C T L 3
C T L 4
Selecting the types of controls to use with this assay
•
Choose one control identifier for each type of control in your assay. The available
options are: Positive Control, Negative Control, High Positive Control, Low Positive
Control, CTL1, CTL2, CTL3, CTL4.
•
After choosing an identifier for Control #1, press ENTER to choose the identifier for
the next control.
Number of Control Replicates
The Number of [Control] Replicates entry screen is presented if the number of control groups is
greater than 0.
N U M B E R
O F
P C
R E P L I C A T E S :
0 2
Entering the number of PC (Positive Control) replicates
•
The well ID associated with Control #1 appears first. Press ENTER to advance to the
next control.
•
Use the NUMERIC keys to enter a value for Number of [Control] Replicates.
•
The valid entry range is from 1 to 12 replicates. The software automatically performs
a check to ensure the number of replicates, multiplied by the number of controls, does
not exceed the number of undefined wells remaining on the plate.
3-26
Operation
Location of Controls
If mapping is manual and controls are defined, the locations for each control replicate must be
specified.
C O N T R O L # 1
L O C A T I O N
T Y P E : P C
R E P # 1 : B 0
1
Entering the control locations (if manual map)
•
Use the NUMERIC and ALPHA keys to enter the well location for Rep #1 of Control
#1. Press ENTER to advance to the next replicate or control group.
Number of Samples
The number of sample groups on the plate can be defined here, and/or it can be defined at run-time
if UTIL
PROMPT FOR SAMPLE COUNT? is set to YES. See Selecting Read
READ
Options in Chapter 2 for more information.
E N T E R
N U M B E R
O F
S A M P L E S : 2 4
Entering the number of sample groups on the plate
•
Use the NUMERIC keys to enter the number of sample groups on the plate.
•
The valid entry range is from 01 to the number of undefined well locations remaining
on the plate.
•
If the Number of Samples setting is altered, the Number of Sample Replicates setting
resets to 01.
Number of Sample Replicates
After the number of sample groups is specified, the Number of Sample Replicates entry screen is
presented.
E N T E R
O F
N U M B E R
S A M P L E
R E P L I C A T E S :
0
2
Entering the Number of Sample Replicates
PowerWaveX Operator’s Manual
3-27
•
Use the NUMERIC keys to enter the number of sample replicates.
•
The valid entry range is from 1 to 12 replicates. The software automatically performs
a check to ensure that the number of replicates multiplied by the number of samples
does not exceed the number of undefined wells remaining on the plate.
Sample Location
If mapping is manual and samples are defined, the locations for each sample replicate must be
specified.
S A M P # 1
L O C A T I O N
R E P # 1 : A 0
1
Defining the locations of the samples
•
Use the NUMERIC and ALPHA keys to enter the well location for Rep #1 of Sample
Group #1. Press ENTER to advance to the next replicate or sample group.
3-28
Operation
Define FORMULA
The PowerWaveX supports four types of formulas (Cutoff, Transformation, Validation, and
General), as well as the ability to program variables for use within formulas. Up to three types of
Validation formulas may be defined (Blank, Control, and Assay Validation).
To define formulas, start at the Main Menu and select DEFINE, select the assay, press ENTER, then
select FORMULA.
D E F I N E
M E T H O D
M A P
F O R M U L A
C U R V
E
Selecting the Formula option on the DEFINE screen
iii
Note: Formulas created using Bio-Tek's Extensions™ (Define Reader Protocol)
software cannot be edited via the reader front panel.
CALCULATION STRUCTURE __________________________________________________
During data reduction, formulas are processed in the order shown below. The number of permitted
formulas of each type are shown as well.
•
Blank Validation
0-1
•
Control Validation
0-4
•
Assay Validation
0-4
•
Transformations
0-1
•
Cutoff Formulas
0-1
•
Curve Fit Analysis (if a curve fit method is defined)
•
General Formulas
PowerWaveX Operator’s Manual
0-4
3-29
FORMULA TYPE ___________________________________________________________
The PowerWaveX supports four types of formulas, as well as the ability to program variables for
use within Transformation formulas.
S E L E C T
F O R M U L A
C U T O F F
T R A N S
S E L E C T
F O R M U L A
G E N E R A L
T Y P E :
V A L
* M O R E
T Y P E :
T R A N S - V A R
Selecting the type of formula to create or edit
•
CUTOFF formulas are used to classify results. During data reduction, results are
evaluated against the cutoff formulas and each well is assigned a user-specified label
(POS, NEG, or EQUIV).
•
TRANSformation formulas are applied to the absorbance data in preparation for
further data reduction and/or curve fit calculation.
•
VALidation formulas can be used to determine whether or not blanks and/or controls
are valid. In addition, Assay Validation formulas can be used to determine whether or
not the entire assay should be considered valid.
•
GENERAL formulas are calculated after all other calculations are complete, and the
results are not used in further data reduction.
Note: General formulas are not currently supported in the PowerWaveX on-board
software.
•
The TRANS-VAR option allows you to define a variable to be used in transformation
formulas.
3-30
Operation
FORMULA ENTRY _________________________________________________________
After the formula type is selected, the Formula Entry screen appears. Each formula can contain a
maximum of 24 characters. Spaces are not necessary.
F O R M U L A # 1 :
M A T H
O T H E R
M A P
F U N C T
N
M A P
F U N C T
N
P C ; x > = 2 . 5 0 0
M A T H
O T H E R
Formula entry screen - the “Formula#1” prompt disappears to provide more spaces
•
After a moment, the FORMULA#1: prompt disappears and the formula can be entered.
Use the options found under MATH, OTHER, MAP, and FUNCTN to “build” the
formula.
To cycle through the available Math, Other, Map, or Function options,
continue to press the appropriate SoftKey. For example, press the MATH
Softkey several times to see +, -, *, /, %, =, etc. When the desired option
appears, press the RIGHT ARROW key to select it and advance to the next
editable field.
Press the LEFT ARROW key to move the cursor to the left.
Press CLEAR to delete the item above the cursor.
When a formula is complete, press ENTER to continue.
•
Select MATH to insert a mathematical symbol such as +, %, or <=.
•
Select OTHER to insert an opening “ ( ” or closing “ ) ” parenthesis, or logical
operators AND or OR.
iii
•
Select MAP to insert a well ID such as BLK;x or NC;1.
•
Select FUNCTN to insert a mathematical function such as LOG or SQRT.
Note: The reader software checks the formulas for errors during data reduction. A
syntax error in a formula will result in a “Token Error” on results reports.
PowerWaveX Operator’s Manual
3-31
MATH
The following mathematical symbols can be used in formulas:
+
Addition
==
Equal to
-
Subtraction
>
Greater than
*
Multiplication
>=
Greater than or equal to
/
Division
<
Less than
%
Percent
<=
Less than or equal to
OTHER
The following additional symbols can be used in formulas:
(
Left parenthesis
)
Right parenthesis
AND
Logical AND
OR
Logical OR
MAP
The available MAP options depend on the formula type and the current plate map.
MAP options resemble BLK;x (mean of the blank wells), NC;1 (the first NC well), or OD (every
well).
FUNCTION
The following functions can be used in formulas:
3-32
LOG10
Log Base 10
ALOG
Anti Log
ALOG10
Anti Log Base 10
LOG
Log
AB
Absolute Value
SQRT
Square Root
PWR
Power
Operation
VALIDATION FORMULAS ____________________________________________________
Validation formulas can be used to determine whether or not blanks and/or controls are valid. In
addition, Assay Validation formulas can be used to determine whether or not the entire assay should
be considered valid.
See Formula Type on page 3-30 for instructions on selecting an assay and accessing the Select
Validation Type screen.
S E L E C T
V A L I D A T I O N
T Y P E :
A S S A Y
B L A N K
C O N T R O L
Selecting a Validation formula type
Control and Blank Validation Formulas
Blank Validation is used to ensure that the OD values for the blank replicates, or for the blank
mean, meet certain criteria. Control Validation serves the same purpose as Blank Validation, but
apply to the control replicates or control mean. If the criteria are not met, results are considered
suspect, and the message “RESULTS INVALID! Blank (or Control) validation failed” appears on
results reports.
•
One blank validation formula can be defined.
•
Up to 4 control validation formulas can be defined.
•
Define the plate map (via DEFINE
MAP) before creating blank or control
validation formulas.
•
Blank/Control validation can be performed on individual replicates (BLK, PC), or on
the group mean (BLK;x, NC;x).
Examples
If an assay protocol states that:
•
Each Blank well should have an OD less than 0.050. The formula is:
BLK<0.050
•
Each Positive Control replicate must fall within the OD range of 1.000 to 2.500. This
can be accomplished with one formula: PC>1.000ANDPC<2.500, or with two
separate formulas: PC>1.000 and PC<2.500
•
The Negative Control mean must have an OD less than 0.100. The formula is:
NC;x<0.100
PowerWaveX Operator’s Manual
3-33
Number of Required Blanks / Controls
When a blank or control validation formula is defined, enter the number of blanks or controls that
must meet the criteria established by that formula.
P C :
N U M B E R
R E P L I C A T E S
O F
V A L I D
R E Q U I R E D :
0 2
Entering the number of blanks/controls that must be found valid
•
Use the NUMERIC keys to enter the number of blanks or controls that are required to
be found valid in order for the results to be valid. For example, if an assay states that
two out of three PC wells must be valid, enter 02.
•
The range is 1 through the number of defined replicates of the blank or control.
Assay Validation Formulas
Assay Validation formula(s) establish a set of criteria used to determine whether or not an assay can
be considered valid. If the criteria are not met, results are considered suspect, and the message
“RESULTS INVALID! Assay validation failed” appears on results reports.
•
Up to four assay validation formulas can be defined.
•
Define the plate map (via DEFINE
MAP) before creating assay validation
formulas.
Examples
If an assay protocol states that in order for the assay to be valid:
•
The mean of the negative controls must be less than 0.100. The formula is:
NC;x<0.100
•
The mean of the positive controls must be greater than the mean of the negative
controls. The formula is:
PC;x>NC;x
3-34
Operation
TRANSFORMATION FORMULAS _______________________________________________
Transformation formulas can be used to transform raw or blanked absorbance data in preparation
for further data reduction, including curve fit analysis.
See Formula Type on page 3-30 for instructions on selecting an assay and accessing the
Transformation Formula definition screen.
•
If a blanking method is selected in the assay, transformation formulas are applied to
the blanked absorbance values, otherwise they are applied to the raw data. Turn to
page
3-29 to review the results calculation structure.
•
One transformation formula may be defined per assay.
•
A transformation formula can be simple (ex. OD*100 to multiply all wells on the plate
by 100), or more complex with the inclusion of a pre-defined Transformation Variable
(see TVAR, below).
Simple Transformation Formulas
“Simple” transformation formulas are typically applied to all wells on the plate. For example:
•
To divide the OD in each well on the plate by 2 and then multiply by 100, the formula
is: (OD/2)*100
•
To raise the OD in each well to the power of 10, the formula is: ODPWR10
Transformation Variable (TVAR)
For more complex transformations, a Transformation Variable (TVAR) can be defined for use
within a transformation formula. This variable defines the scope of the transformation: whether to
apply the transformation to all of the wells on the plate (OD), or to just the sample wells (SMP).
S C O P E
V A R I A B L E :
S M P
O D
O D
Choosing the scope of the transformation
PowerWaveX Operator’s Manual
3-35
•
If SMP is chosen:
The transformation formula will be applied to the sample wells only.
SMP and any other well identifiers (BLK, PC, NC, STD, etc.) defined will
become available as MAP options when building the transformation formula.
Example:
The assay plate map contains 2 NC wells and 2 PC wells. The remainder of
the map is filled with samples.
The assay data reduction requires that the mean of the NC be subtracted from
all the samples on the plate.
The transformation formula is: SMP-NC;x
•
If OD is chosen, the formula definition screen will appear so that you can define a
formula for use within the transformation formula.
Use the formula keys (Math, Other, Map and Function) to define the
Transformation Variable (TVAR). Once the variable has been defined, it can
be used in a transformation formula. The TVAR will be available as a MAP
option when building the transformation formula.
Example:
The assay plate map has 2 blanks, 1 control well in duplicate (CTL1), 1 negative
control well in triplicate (NC), and 5 standards in duplicate (STD1-STD5) in varying
concentrations.
The assay data reduction states:
Subtract the mean of CTL1 from the mean of the NC. Subtract the difference
from all OD’s on the plate.
Divide the result of the above by the mean of the NC less the mean of CTL1,
then multiply by 100.
On paper, the formula reads:
(OD - (NC;x-CTL1;x)) / (NC;x-CTL1;x) * 100
On the reader, the formula (NC;x-CTL1;x) will be programmed as the TVAR,
since the transformation will apply to all standards, controls and samples on the plate.
To do this:
At the Scope Variable selection screen, choose OD and press ENTER.
Enter the formula (NC;x-CTL1;x) using the Math, Other, Map and
Function keys. Press ENTER.
3-36
Operation
The formula selection screen is displayed. Choose TRANS.
Enter the formula (OD-(TVAR))/(TVAR)*100 using the Math, Other,
Map and Function keys. “TVAR” is available as a MAP option.
Example:
In the case of competitive reactions, converting absorbance data to percent B/B0 can
be: (OD/STD1)*100. This divides all the wells by STD1, presumably the 0
standard, and multiplies the results by 100. To do this:
At the Scope Variable selection screen, choose OD and press ENTER.
Enter simply STD1 as the TVAR formula. Press ENTER.
The formula selection screen is displayed. Choose TRANS.
Enter the formula (OD/TVAR)*100 using the Math, Other, Map and
Function keys. “TVAR” is available as a MAP option.
PowerWaveX Operator’s Manual
3-37
CUTOFF FORMULAS ________________________________________________________
A cutoff formula calculates a cutoff value that is used for classifying samples. See Formula Type
on page 3-30 for instructions on selecting an assay and accessing the Cutoff formula definition
screen.
During data reduction, results are evaluated against the cutoff value (with an optional greyzone),
and each well is assigned a call POS (positive), NEG (negative), or EQUIV (equivocal).
•
One cutoff formula may be defined per assay.
•
If Transformation Formulas are defined, cutoffs are based on the transformed results.
Turn to page 3-29 to review the results calculation structure.
•
A cutoff formula can consist of a simple numeric value (1.500), a well identifier (PC,
or PC;x to represent the mean), or a formula combining the two (NC;x+0.050).
•
A “greyzone” around the cutoff value can be defined, to indicate questionable results.
•
Do not use the < or > mathematical symbols in a cutoff formula.
•
Tip: Choose to print a Column Report to see the greyzone and cutoff values,
as well as the equations used to assign calls to samples. See Specifying Data Output
and Reporting Options in Chapter 2 for information on Column Reports.
Greyzone
The greyzone is a definable area around the cutoff value. Samples falling within an area defined
by the greyzone (ex. ± 5% of the cutoff value) could be considered questionable, or equivocal
(EQUIV).
E N T E R
G R E Y Z O N E :
0 5 %
Defining a greyzone around the cutoff value
•
Use the NUMERIC keys to enter the greyzone percentage.
•
The valid entry range is from 00 to 99%. An entry of 00% indicates no greyzone,
although a sample equal to the cutoff value will still receive the EQUIV call.
•
3-38
See POS / NEG Calls on the next page for information on how calls are assigned.
Operation
POS / NEG Calls
After the greyzone is defined, calls for the sample wells (POSitive, NEGative, EQUIVocal), must be
defined.
S A M P > C U T O F F + 1 5 % :
P O S
P O S
N E G
Defining the POS/NEG/EQUIV calls for samples
•
Select POS or NEG to select the call that will be assigned to samples greater than the
cutoff value plus the greyzone.
•
If for example POS is selected in the screen shown above, calls will be assigned
according to the following equations (SMP represents the sample wells):
EQUIV:
SMP <= (CUTOFF+(CUTOFF*GREYZONE)) AND
SMP >= (CUTOFF-(CUTOFF*GREYZONE))
POS:
SMP >
(CUTOFF+(CUTOFF*GREYZONE))
NEG:
SMP <
(CUTOFF-(CUTOFF*GREYZONE))
Examples
1.
The cutoff between negative and positive calls should be calculated as the average of the
negative controls plus the OD value 0.050. Samples greater than the cutoff should be
labeled as positive. No greyzone is required.
•
For this example, NC;x (the mean of the NC wells) equals 0.100 OD
•
The cutoff formula is NC;x+0.050
•
The greyzone is 00%
•
POS is selected for SAMP>CUTOFF+00%
•
Calls are assigned to sample wells as follows:
EQUIV if the sample equals 0.150
POS if the sample is greater than 0.150
NEG if the sample is less than 0.150
PowerWaveX Operator’s Manual
3-39
2.
For a quantitative assay, samples with ODs greater than the STD2 mean plus a 10%
greyzone should be labeled as positive, samples with ODs less than the STD2 mean minus
the 10% greyzone should be labeled as negative. All other samples should be considered
equivocal.
•
For this example, STD2;x (the mean of the STD2 wells) equals 1.500 OD
•
The cutoff formula is simply STD2;x
•
The greyzone is 10%
•
POS is selected for SAMP>CUTOFF+10%
•
Calls are assigned to sample wells as follows:
EQUIV if the sample is greater than or equal to 1.350 and less than or equal
to 1.650
POS if the sample is greater than 1.650
NEG if the sample is less than 1.350
GENERAL FORMULAS ______________________________________________________
General Formulas are calculated after all other calculations are complete, and the results are not
used in further data reduction.
Note: General Formulas are not supported in the current version of the reader.
3-40
Operation
Define CURVE
To define curve-fitting parameters for an assay, start at the Main Menu and select DEFINE, select
the assay, press ENTER, then select CURVE.
D E F I N E
M E T H O D
M A P
F O R M U L A
C U R V E
Selecting the Curve option on the DEFINE screen
Configurable curve-fitting parameters include:
•
Curve-Fit Type
•
Editing of Outliers
•
Axis Identification
•
Extrapolation of Unknowns
CURVE-FIT TYPE __________________________________________________________
The PowerWaveX supports 7 different curve-fitting methods, Linear, Quadratic, Cubic, 4-P, 2-P,
cubic-spline, and point-to-point.
C U R V E - F I T
N O N E
T Y P E :
L I N E A R
C U R V E - F I T
C U B I C
C U R V E - F I T
C - S P L I N E
Q U A D
T Y P E :
4 - P
L I N E A R
L I N E A R
2 - P
T Y P E :
* M O R E
* M O R E
L I N E A R
P T - P T
* M O R E
Selecting a curve-fit type
•
NONE: Choose None if no standard curve will be generated (this is the default).
•
LINEAR: A simple best fit straight line is plotted using the values of the standards.
PowerWaveX Operator’s Manual
3-41
•
2
QUADratic: Uses the Quadratic equation “ax +bx +c=y” to plot the standards values.
Utilizing this curve, any data point for a standard that deviates from the ideal value
will not affect the entire curve.
•
CUBIC: Uses the equation “ax3 + bx2 + cx + d = y” to plot the standards values. This
type of curve fit is affected even less than the quadratic fit when any particular
standard has a poor value.
•
2-P (Logit/Log): A curve fitted to the standard values, which is characterized by a
skewed sigmoidal (S-shaped) plot that eventually becomes asymptotic to the upper
and lower standard values. The logistic equation is algebraically transformed to a
simpler form in which experimentally determined values are used for the responses at
concentrations of zero and infinity.
•
C-SPLINE (Cubic-Spline): A piecewise polynomial approximation consisting of
joining a set of data points by a series of straight lines, which is then smoothed by
using a cubic fit.
•
4-P (4-Parameter Logistic): A curve fitted to the standard values, which is
characterized by a skewed sigmoidal (S-shaped) plot that eventually becomes
asymptotic to the upper and lower standard values. The 4 parameters are: Left
asymptote, Right asymptote, Slope and Value at the Inflection point. This fit is most
recommended for immunoassay data, and is more exact than Logit/Log.
•
PT-PT (Point-to-Point): A plot that connects each standard point with a line, with no
averaging of the values to “smooth” the curve at each standard.
EDIT STANDARD OUTLIERS __________________________________________________
After the standard curve has been generated, one or more standards can be excluded from the
recalculation of the curve.
E D I T
S T D
N O N E
O U T L I E R S : M A N U A
L
M A N U A L
Choosing whether or not to enable standard outlier editing for this assay
•
Select NONE to suppress the Edit Standard Outliers capability for this assay.
•
Choose MANUAL to enable the capability.
If AVERAGE STANDARDS is set to NO, the individual standard replicates are
available for editing. If set to YES, the standard groups are available for
editing.
3-42
Operation
After the assay is run and reports are generated, press REPORT from the
Main Menu. Press RESULT, select the assay, then press ENTER. The EDIT
STD OUTLIERS? YES/NO prompt will appear. See Editing Standard
Outliers on page 3-53 for further instructions.
X/Y AXIS TYPE ___________________________________________________________
After the curve-fit type is selected, select the X/Y Axis Type.
X / Y
L I N
A X I S
T Y P E :
L I N / L O G
L I N
L O G
L O G / L I N
Selecting the X/Y Axis Type
•
Choose the method by which the X and Y axes will be scaled.
•
This option is not available for the 2-P and 4-P curve-fit types. The X/Y scaling for
these curves is always LIN/LIN.
EXTRAPOLATION OF UNKNOWNS _____________________________________________
The PowerWaveX provides the option to “extrapolate” the curve to evaluate samples outside of the
absorbance range defined by the standards.
E X T R A P O L A T E
Y E S
U N K N O W N S ? Y E
S
N O
Choosing whether or not to extrapolate unknowns
•
Select YES to enable Extrapolation, otherwise select NO.
•
On printed reports, extrapolated concentrations (RSLT values) are surrounded by < >,
for example <44.425>.
iii
Note: If extrapolation is chosen for the Point-to-Point curve fit, unknown
concentrations will be extrapolated linearly from the nearest segment of the curve. If
the plot includes both increasing and decreasing segments, the curve printout will be
labeled “Ambiguous.” The resulting values, which actually are extrapolated, may not
be indicated as such. All calculated results for an “Ambiguous” curve should be
considered unreliable.
PowerWaveX Operator’s Manual
3-43
Panel Assays
A Panel assay is a collection of up to eight assays to be run on one plate.
•
The most common reason to use a Panel assay is for confirmatory tests based on a
screening test in clinical applications.
•
Only one panel assay can be defined on the reader at any time.
•
The assays specified within the Panel must be pre-defined in any of the assay positions
1-55.
•
The assays specified within the panel must all use the Endpoint read method.
•
The assays specified within the panel must all read at the same wavelength(s).
•
Any curve-fit type, formulas or standard concentrations previously defined for each
assay will be used when the assay is selected for a Panel.
•
The type and number of controls, blanks, standards and replicates in the assays chosen
for the Panel will be “copied” into the Panel definition. To change any of the map or
assay parameters in the Panel, they must be changed in the pre-defined assay first.
•
Consider printing a Map Report for each assay that will be included in the panel, for
use with mapping the Panel.
To create a panel assay, start at the Main Menu, press DEFINE, then choose assay number 99.
Enter the panel assay name.
N A M E :
-
P A N E L
/
:
S P A C E
Editing the name of the Panel assay
3-44
•
The default name is “PANEL”.
•
Use the ALPHA and NUMERIC keys to update the Assay name, if desired.
•
Press ENTER to continue. The Number of Assays entry screen will appear.
Operation
N U M B E R
O F
A S S A Y S :
2
Entering the number of assays to include in the Panel
•
Specify the number of assays to include in the panel (1 to 8).
•
Press ENTER to continue. The Mapping Direction selection screen will appear.
M A P P I N G
D O W N
D I R E C T I O N : D O W N
A C R O S S
Choosing the Panel's mapping direction
•
This option ensures that all assays will be mapped in the same direction. Select DOWN
or ACROSS.
iii
Note: The original map directions for the pre-defined assays are overridden by the
Panel's direction. If the assays include replicates, they will follow the Panel map
direction.
After selecting the mapping direction of the assays, choose which assays to include in the panel.
S E L E C T
N A M E :
A S S A Y
N U M B E R :
2 2
H B S - A G 1
Selecting the assays to include in the panel
•
Press OPTION to cycle through the assay numbers and names, or use the NUMERIC
keys to enter an assay number. Press ENTER to make a selection.
•
After selecting an assay, its starting location must be defined.
PowerWaveX Operator’s Manual
3-45
S T A R T
A T
M A P P I N G
W E L L
L O C A T I O N :
A 0
1
Defining the starting location for the selected assay
•
Use the ALPHA and NUMERIC keys to choose the well location to begin the assay.
Wells A01 through H01 are valid for ACROSS mapping; A01 through A12 are valid
for DOWN mapping.
•
This process will be repeated for each assay within the panel. Remain aware of the
total number of controls, standards and blanks that were originally mapped in each
individual assay while mapping for the Panel assay.
For example, to include Assays 1, 8 and 22 in the Panel assay (DOWN mapping is
selected for the Panel):
Assay 1 has a total of 12 wells defined for controls, blanks and standards. In
the Panel, the mapping for Assay 1 begins in well A01. The user wants to run
6 samples in Assay 1. Assay 1 now fills wells A01 through B03.
The mapping for Assay 8 can begin in well B04, or any well other than A01
to B03. The reader will “beep” if you try to map into a well that is already
assigned for use with the Panel.
The mapping for Assay 22 may begin at the next available well location after
Assay 8 mapping is complete.
After all the assays have been entered into the Panel, consider printing the
Panel's Map Report to verify the map before reading the plate. Choose
REPORT (from the Main Menu), MAP, ASSAY 99. The reader will print the
map of each assay configured in the Panel.
The Panel Assay results are sorted by Sample (unless a custom assay has
been programmed by Bio-Tek).
iii
Note: The Interpretation of Results reports for each assay in the panel will
print first, and then the Sample results will print. See Appendix D for a
sample Panel Report.
3-46
Operation
Reading a Microplate
Use the Main Menu option READ, or press the READ key on the front panel, to initiate a plate read.
(MAIN)
READ
DEFINE
REPORT
UTIL
↓
SELECT ASSAY NUMBER: 05
NAME: DNA Quant
Press OPTIONS to scroll through the assay list.
Press ENTER to select an assay.
↓
ENTER NUMBER OF
SAMPLES:
45
↓
PLATE ID: 45768-A
-
/
:
SPACE
↓
ENTER
SAMPLE ID: 0001
↓
PLACE PLATE IN CARRIER
AND PRESS <READ> KEY
Figure 3-5: Options available under READ
iii
Notes: The options to present the Enter Number of Samples, Plate ID, and/or Sample
ID screens are configurable via UTIL
READ. Custom assays may present
additional screens for the entry of special parameters; refer to the Assay Reference
Guide (PN 7271006) for more information.
iii
Notes: Before reading a plate, ensure that the reporting options are set correctly under
UTIL
OUTPUT. See Selecting Read Options in Chapter 2 for more information.
PowerWaveX Operator’s Manual
3-47
SELECTING AN ASSAY ______________________________________________________
From the Main Menu, press READ and then select the desired assay.
S E L E C T
A S S A Y
N U M B E R : 6 5
N A M E : H B S - A G 1
Selecting an assay to run
•
Use the NUMERIC keys to enter the number of any predefined assay stored in the
reader’s memory, or the OPTION key to advance one assay at a time.
•
Press ENTER to continue.
RUN-TIME PROMPTS _______________________________________________________
After the assay is selected, one or more informational prompts may be presented, depending on
preferences selected via UTIL
READ, whether or not the assay specifies manual mapping, or if
this is a custom pre-programmed assay.
•
Prompts enabled via UTIL
READ can include Enter Number of Samples, Plate
ID, and Enter Sample ID.
•
If the assay specifies manual mapping, prompts for information will include the
locations for the sample wells.
•
If running a custom assay, typical prompts might include (refer to the Assay Reference
Guide for more information):
The number of samples
Standard concentrations
Assay ID
Fill pattern
Blank method
First well location
Replicate count for each well type
Wavelength mode
Report preferences, etc.
3-48
Operation
Enter Number of Samples
If the Enter Number of Samples prompt is presented, indicate the number of sample groups on the
plate. The number of sample replicates is typically pre-defined in the assay, but if this is a custom
assay, you may also be prompted to enter the replicate count.
E N T E R
N U M B E R
O F
S A M P L E S :
2 0
Entering the number of sample groups on the plate
•
Use the NUMERIC keys to enter the number of sample groups.
•
The valid entry range is from 01 to the maximum number of wells remaining on the
plate after any blank, control, or standard wells are mapped.
•
If you enter a value greater than the number of empty wells remaining on the plate, the
reader will “beep” and automatically change the value to the maximum permissible
number of samples.
Enter Plate ID
If the Plate ID prompt is presented, enter a unique plate identifier to be stored in memory with the
assay name and absorbance data.
P L A T E
-
I D :
0 0 1 - A
/
:
S P A C E
Entering a Plate ID
•
Use up to 10 alphanumeric characters. See Figure 3-1 for instructions on using the
keypad.
•
Although the reader does not require it, consider entering a unique ID for each plate,
especially if reading multiple plates for the same assay.
•
If the internal bar code scanner option is installed, the reader will automatically scan
the plate/bar code label and use that for the Plate ID.
iii
Note: Use caution when creating multiple Plate IDs. The reader does not provide a
warning when the maximum of 8 Plate IDs stored in memory is about to be exceeded.
If a 9th Plate ID is added, it will overwrite the first Plate ID stored in memory.
PowerWaveX Operator’s Manual
3-49
Enter Sample ID
If the Enter Sample ID prompt is presented, enter a starting sample identification number.
E N T E R
S A M P L E
I D :
0 0 0 1
Entering the starting sample identification number
•
The valid entry range is from 0001 to 9999.
•
The software will automatically increment each subsequent sample identification by 1.
•
The sample IDs will be assigned in accordance with the plate map defined in the
assay.
Enter Well Location
If the assay specifies manual plate mapping and if Prompt for Sample Count is set to Yes under
READ, sample well locations can be defined at run-time.
UTIL
S A M P L E #
1
L O C A T I O N
R E P #
1 : A 0 2
Entering the sample well locations at run-time
•
The sample well locations originally defined in the assay will be presented.
If desired, use the keypad to enter new well location for each sample replicate.
•
3-50
Press ENTER to advance to the next replicate.
Operation
BEGINNING THE PLATE READ ________________________________________________
When all required prompts have been responded to, the Place Plate in Carrier prompt appears.
P L A C E
A N D
P L A T E
P R E S S
I N
C A R R I E R
< R E A D >
K E Y
Final prompt before the plate read begins
•
Before reading the plate, make sure that the printer is connected, turned on, and full of
paper.
•
Place the plate on the carrier, then press the READ key to initiate the plate read.
•
After the read is complete, data reduction will perform (“Calculating Results…”), and
then the reports will be generated (“Generating Reports…”).
•
iii
To halt in read in progress, press the STOP key.
Note: If using the incubation option, the reader will wait for the incubator to reach
temperature before reading the plate.
PowerWaveX Operator’s Manual
3-51
Printing Reports
Reports are automatically generated after a plate has been read (see Specifying Data Output and
Reporting Options in Chapter 2 for information on selecting reports). Results reports also can be
regenerated manually by using the REPORT option from the Main Menu. In addition, Map, Assay,
and Assay List reports can be printed.
Note: See Appendix D for sample reports.
R E A D Y
9 : 4 5 A M
R E A D
D E F I N E
P R I N T
R E P O R T ?
R E S U L T
M A P
0 7 / 2 0 / 9
R E P O R T
A S S A Y
U T I
9
L
L I S T
Generating reports via the REPORT option from the Main Menu
•
Select RESULT to obtain an exact copy of results from the plate reading (the 8 most
recent sets of plate data are stored in memory).
The form in which the results are presented is determined by the report
settings (Matrix, Column, Curve Fit) specified under UTIL
•
OUTPUT.
Select MAP to print a matrix showing the locations of the Blanks, Standards, Controls
and Samples for a particular assay.
•
Select ASSAY to print a plate map and a listing of all of an assay’s settings, such as
wavelengths, numbers of well types, formulas, and curve fit parameters.
•
Select LIST to print a list of all assays (name and number), currently programmed in
the reader.
3-52
Operation
Results Report
The reader stores the absorbance data for the 8 most recent plate reads. Results reports can be
generated for these plates if, for example, the data that automatically printed after the read needs to
be printed in a different format, or if the standard curve contains outliers that require editing.
R E P O R T : H B S - A G
I D :
0 0 0 1
0 7 / 2 2 / 9 9
Choosing a plate for the Results Report
•
The most recently read plate is presented first, showing the assay name, the plate ID
(if one was entered), and the date the plate was read.
•
Press OPTIONS to see the next plate in memory.
•
Press ENTER to select a plate and continue.
•
If a standard curve was generated and EDIT STANDARD OUTLIERS was set to
MANUAL in the assay definition, the EDIT STD OUTLIERS? prompt is presented;
otherwise, the Print Results? prompt is presented.
Editing Standard Outliers
If a standard curve was generated and if EDIT STANDARD OUTLIERS was set to MANUAL in the
assay definition, the option to edit outliers is presented.
E D I T
S T D
Y E S
N O
O U T L I E R S ?
Choosing whether or not to edit standard outliers
•
Select NO to include all standards in the curve fit calculations.
•
Select YES to indicate that one or more standard replicates or groups should be
temporarily excluded from curve fit calculations.
If AVERAGE STANDARDS was set to NO in the assay definition, one or more
standard replicates can be chosen for exclusion.
PowerWaveX Operator’s Manual
3-53
E D I T
S T D
Y E S
N O
1
R E P 1 ?
Y E S
Choosing to remove replicate 1 of Standard 1 from curve fit calculations
−
Select YES to exclude the replicate from curve fit calculations.
−
Select NO to retain the replicate.
−
Press ENTER to advance to the next replicate.
If AVERAGE STANDARDS was set to YES in the assay definition, one or
more standard groups can be chosen for exclusion.
E D I T
S T D
Y E S
N O
1 ; x ?
Y E S
Choosing to remove all replicates of Standard 1 from curve fit calculations
iii
−
Select YES to exclude the group from curve fit calculations.
−
Select NO to retain the group.
−
Press ENTER to advance to the next group.
Note: Each curve-fit type requires a minimum number of standards for curve
generation: 4 for 2-P, 4-P, cubic, and cubic-spline, 3 for quadratic, and 2 for linear
and point-to-point. Exercise caution when editing outliers. If the assay is left with
insufficient standards, the curve fit will fail.
Printing Results
After the assay is selected and standard outliers are edited (if necessary), the results report can be
printed.
P R I N T
Y E S
R E S U L T S ?
N O
Print Results screen
3-54
•
Ensure that the printer is connected, turned on, and full of paper.
•
Press YES to print reports, or NO to return to the Main Menu.
Operation
Map Report
The Map Report contains a matrix in Row x Column format, showing the location of every well
identifier defined in the plate map.
S E L E C T
A S S A Y
N A M E :
R N A
N U M B E R :
0 7
Q u a n t
Choosing an assay for the Map Report
•
Press OPTIONS to cycle through the list of available assays, or enter the number of the
desired assay.
•
Press ENTER to print the report.
Assay Report
The Assay Report lists the assay definition parameters and their current settings.
S E L E C T
N A M E :
A S S A Y
N U M B E R : 0 1
H B S - A G
Selecting an assay for the Assay Report
•
Press OPTIONS to cycle through the list of available assays, or enter the number of the
desired assay.
•
Press ENTER to print the report.
List Report
The List Report lists the all of the assays (name and number), currently programmed on the reader.
•
Select REPORT from the Main Menu, then select LIST to print the report.
PowerWaveX Operator’s Manual
3-55
3-56
Operation
Chapter 4
Performance Verification/
Qualification Tests
This chapter discusses the tasks and procedures necessary
for verifying and qualifying instrument performance on an
ongoing basis. A convenient Recommended Test Schedule
arranges tasks into Installation, Performance, and
Operational Qualification categories.
Recommendations for Achieving Optimum Performance
•
Microplates should be perfectly clean and free of dust or bottom scratches. Use new
microplates from sealed packages. Do not allow dust to settle on the surface of the solution; use
microplate covers when not reading the plate. Filter solutions to remove particulates that could
cause erroneous readings.
•
Although the PowerWaveX supports most flat, U-bottom, and V-bottom microplates, the reader
achieves optimum performance with optically clear, flat-bottomed wells.
•
Non-uniformity in the optical density of the well bottoms can cause loss of accuracy, especially
with U- and V-bottom polyvinyl microplates. Check for this by reading an empty microplate.
Dual wavelength readings can eliminate this problem, or bring the variation in density readings
to within acceptable limits for most measurements.
•
Inaccuracy in pipetting has a large effect on measurements, especially if smaller volumes of
liquid are used. For best results, use at least 100 µl per well in a 96-well plate and 25 Ol in a
384-well plate.
•
Dispensing solution into 384-well plates often traps air bubbles in the wells. Dual wavelength
reads will cancel most of these errors; however, for best results, they should be removed by
degassing the plate in a vacuum chamber prior to reading.
•
The inclination of the meniscus can cause loss of accuracy in some solutions, especially with
small volumes. Agitate the microplate before reading to help bring this problem within
acceptable limits. Use Tween® 20, if possible (or some other wetting agent) to normalize the
meniscus. Some solutions develop menisci over a period of several minutes. This effect varies
with the brand of microplate and the solution composition. As the center of the meniscus drops
and shortens the light path, the density readings change. The meniscus shape will stabilize over
time.
Recommended Test Schedule
The schedule shown in Table 4-1 defines the factory-recommended intervals for performance
testing for a microplate reader used for one shift seven days a week.
Note:
The risk factors associated with your tests may require that the Performance and Operation
Qualification procedures be performed more or less frequently than shown below.
Table 4-1
Recommended Test Schedule
Installation
Qualification
Performance Performance
Operation
Qualification / Qualification / Qualification /
Monthly
Quarterly
Annually
System Self-Test, p. 4-4
Universal Plate Test, p. 4-7
Liquid Test 1, p. 4-17
Liquid Test 2, p. 4-19
*
*
*
Liquid Test 3, p. 4-22
Optional for 340 nm
Robotic Lube, p. 4-25
Every six months, or after 10,000 cycles.
* Run Liquid Test 2 only if you do not have a Universal Test Plate. If you run Liquid Test 2 or 3,
you do not have to also run Liquid Test 1.
Installation and Performance Qualification Procedures
Tests outlined in this section may be utilized to confirm initial and ongoing performance of the
PowerWaveX.
Your PowerWaveX reader was fully tested at Bio-Tek prior to shipment and should operate properly
upon initial setup. If it is suspected that problems may have occurred during shipment, if you
reshipped the device, or if regulatory requirements dictate that Performance Qualification Testing is
necessary, the following tests should be performed. After the initial confirmation of operation, the
Universal Plate Test and Liquid Testing should be performed at least monthly. The System Self Test
and Universal Plate Test can be run from KCjunior via Utilities, Diagnostics, and the Universal
Plate Test can be run from KC4 via System, Diagnostics.
•
System Self Test: Verifies proper gains, bulb operation, low electronic noise, and
optional incubator functionality. Test results are sent to an attached printer.
•
Universal Plate Test: Confirms the optical accuracy, linearity, alignment, channel-to-
channel variation, and wavelength accuracy of the instrument
•
Liquid Testing: Quantifies the channel-to-channel variation of the instrument using
liquids, which verifies operation in a way that glass test filters cannot.
4-2
Performance Verification/Qualification Tests
Routine Procedure
To ensure proper operation of the PowerWaveX on an ongoing basis, the System Self-Test and the
Universal Plate Test should be conducted monthly.
•
Select Reader System Test to verify that the light levels and electronic noise at all set
wavelengths fall within factory acceptance criteria.
•
Select Universal Test Plate to run the calibration plate test to confirm the alignment,
repeatability, and accuracy/linearity of the reader. The report will also contain the part
and version numbers of the software loaded on the unit.
•
Perform a wavelength scan to confirm wavelength accuracy of the reader.
From the Main Menu, press the soft key that corresponds to UTIL to access the Utility Options
Menu. The Select Utility Option screen will appear.
R E A D Y
R E A D
9 : 4 5 A M
D E F I N E
S E L E C T
T E S T S
R E P O R T
U T I L I T Y
S E T U P
0 5 / 0 9 / 9
U T I
9
L
O P T I O N :
O U T P U T
R E A D
Selecting UTIL from the Main Menu
•
Select TEST to advance to the Select Test screen.
S E L E C T
T E S T :
S Y S T E M
C H K S U M
C A L P L A T E
Choosing a performance test to run
•
Select SYSTEM to verify that the light levels and electronic noise at all wavelengths
fall within factory acceptance criteria.
•
Select CHKSUM to instruct the reader to compare the software to the internally
recorded checksum values to ensure that the programming has not been corrupted. In
addition, use this option to view the part numbers and versions for the software
currently installed on the reader. This information is needed when contacting Bio-Tek
for technical assistance.
•
Select CALPLATE to run the calibration plate test, to confirm the alignment,
repeatability, and accuracy/linearity of the reader.
PowerWaveX Operator’s Manual
4-3
SYSTEM TEST __________________________________________________________
The System Test confirms that the light levels and electronic noise at all wavelengths fall within
factory acceptance criteria, and accomplishes this by measuring the air and dark readings and
evaluating them to ensure they fall within specified ranges.
The reader automatically runs an internal System Test each time it is powered on. An error will be
displayed if the power on System Test fails. No report will be produced at the power on System
Test.
To obtain a report of the System Test values for either periodic testing documentation or
troubleshooting (Figure 4-1), start at the Main Menu and press UTIL
TESTS
SYSTEM.
The PowerWaveX will conduct the instrument’s System Test and report results in a pass/fail format.
See Appendix C for a list of possible error codes.
Photodiodes
The Optics portion of the System Self Test confirms that the eight reading and one reference
channels have adequate signal range without saturating the electronics.
Xenon Flash Lamp
The Optics test also indicates if the xenon flash lamp is within operational limits.
Incubation
If the PowerWaveX has the 4-Zone incubation option installed, the System Self-Test will verify
the four thermistors and compare these readings to internal voltage references to confirm proper
operation.
4-4
Performance Verification/Qualification Tests
Figure 4-1: Sample Output for the System Test
PowerWaveX Operator’s Manual
4-5
CHECKSUM TEST (CHKSUM)______________________________________________
The Checksum test runs automatically when the reader is powered on. The test compares the
software to the internally recorded checksum values to ensure that the programming has not been
corrupted. If there are any errors during the power-on checksum test, they will be displayed.
To verify the checksum manually, and to view the part numbers and versions of software currently
loaded onto your reader, start at the Main Menu and press UTIL
TESTS
CHKSUM. The
information displayed will resemble the following; the actual checksum will be dependent upon the
software version that is loaded:
7 2 6 0 2 0 1
C o d e
V e r s i o n
C h e c k s u m :
1 . 1 9
( 0 8 5 D )
The initial checksum test display, showing the basecode software part number, version number,
and checksum. After a few moments, a second screen will display.
7 2 6 0 2 1 2 - F W
v 2 . 0 0 . 0 0
The second checksum test display, showing the assay configuration software part number and
version number.
4-6
Performance Verification/Qualification Tests
Calibration Verification
It is considered good laboratory practice to periodically verify the calibration of the PowerWaveX.
Verification should be performed monthly using the tests in this section:
•
Universal Plate Test (see below)
•
Liquid Tests (see page 4-14)
UNIVERSAL PLATE TEST __________________________________________________
The Universal Plate Test (also referred to at the Calibration Plate Test), confirms the alignment,
repeatability, and accuracy/linearity of the PowerWaveX. An alternative method used to determine
accuracy, repeatability and linearity is the Liquid Test 2 described later in this chapter.
The Universal Test Plate (PN 7260522) allows comparison of the reader’s optical density
measurements and mechanical alignment to NIST traceable values. Accuracy/ linearity,
repeatability, and alignment are tested. Specific standard calibration values must be entered for
each wavelength to be tested (see Entering the Universal Test Plata Data on the following page).
The Universal Plate Test confirms the following:
•
Accuracy of the Optical Density readings -- the comparison of the optical density
readings with those given with the Universal Test Plate insert will confirm the
accuracy of the optical density readings at specific wavelengths.
•
Linearity of the Optical Density readings are confirmed by default if the optical
density readings are accurate. This can be proven by analyzing the data in a program
such as Microsoft® Excel.
•
Alignment of the plate carrier and standard microplates are confirmed by the four
corner positional accuracy check.
•
Channel-to-channel variation can be tested by completing the turnaround test.
This tests the reader’s ability to read samples accurately in different plate positions.
•
Wavelength setting accuracy. To check the accuracy of wavelength settings, use
the Universal Test Plate. The Test Plate provides a multiband test filter (Didymium
glass V10) in location C6.
PowerWaveX Operator’s Manual
4-7
Requirements
To run the Universal Plate Test from the reader front panel, you need Bio-Tek's Universal
7-Filter Test Plate (PN 7260522), with its accompanying Data Sheet, shown in Figure 4-2 below.
Figure 4-2: Sample Universal Test Plate data sheet
Entering the Universal Test Plate Data
iii
Note: Before defining the standard calibration values, set the reader's internal
wavelength settings to correspond with up to six of the wavelengths shown on the
Universal Test Plate Data Sheet (see Figure 4-2 above). See Adjusting the Reader's
Wavelength Table Settings in Chapter 2 for instructions. Additional wavelengths,
above 405 nm, are available at extra cost. Contact your Bio-Tek dealer.
Refer to the Data Sheet provided with the Universal Test Plate (Figure 4-2), when entering the
standard values. To enter the values into the reader, start at the Main Menu and press UTIL
SETUP. The Edit Setup Information menu will appear.
E D I T
S E T U P
D A T E
E D I T
T I M E
S E T U P
R S 2 3 2
C A L
I N F O R M A T I O N :
L A M B D A
* M O R
E
I N F O R M A T I O N :
P L A T E
* M O R
E
Edit Setup Information menu. Press *MORE to see the Cal Plate option.
•
4-8
From the Edit Setup Information menu, press *MORE, then select CAL PLATE.
Performance Verification/Qualification Tests
The Calibration Lambda entry screen will appear.
C A L I B R A T I O N
4 0 5
4 5 0
L A M B D A :
4 9 0
4 0 5
6 3 0
Selecting a wavelength for which to enter standard calibration values
•
Choose a wavelength (e.g., 405), then press ENTER.
W A V E L E N G T H : 4 0 5
C A L I B R A T I O N
W E L L : C 0
V A L U E S
0 . 1 4
1
7
Entering standard calibration values, referencing the Universal Test Plate Data Sheet
•
Enter the absorbance value from the Data Sheet for the selected wavelength and
current well location (C01 in the above example).
•
After each entry, press ENTER to advance to the next consecutive well location.
•
When the sixth value has been entered, press ENTER to return to the Calibration
Lambda selection screen to select another wavelength, or Main Menu.
Running the Universal Plate Test
Before running the Universal Plate Test, ensure that the standard calibration values for the test plate
have been defined on the reader (see Entering the Universal Test Plate Data on the previous page).
To run the Universal Plate Test:
•
Ensure that the printer is attached, turned on, and full of paper.
•
From the reader's Main Menu, press UTIL
TESTS
CAL PLATE.
The Calibration Lambda selection screen will appear.
C A L I B R A T I O N
4 0 5
4 5 0
L A M B D A :
4 9 0
4 0 5
6 3 0
Selecting a wavelength for the calibration plate test
•
Select a wavelength for the test, then press ENTER.
PowerWaveX Operator’s Manual
4-9
The Place Plate in Carrier prompt will appear.
P L A C E
A N D
P L A T E
P R E S S
I N
C A R R I E R
< R E A D >
K E Y
Place the Universal Test Plate in the reader, then press READ
•
Place the Universal Test Plate in the carrier so that well A1 is in the upper left corner
of the carrier, then press READ to begin the test.
The plate will be read twice, then the Rotate Plate prompt will appear.
R o t a t e
P r e s s
P l a t e
< R E A D >
1 8 0
t o
d e g r e e s
C o n t i n u e
Rotate the plate, then press READ
•
Rotate the plate so that well A1 is in the lower right corner of the carrier, then press
READ to complete the test.
•
When the test is complete, results will print. See the sample Calibration Plate
Analysis report in Figure 4-3.
4-10
Performance Verification/Qualification Tests
Figure 4-3: Sample printout showing the calibration plate analysis
The Calibration Plate Analysis Report contains results for the following:
•
Alignment: This portion of the test measures the alignment of the microplate carrier
with the optical path. A reading > 0.015 represents an out-of-alignment condition.
Wells A01, A12, H01, and H12 are the only valid alignment holes for the reader on
the PN 7260522 Calibration Test Plate.
•
Accuracy: Accuracy is a measure of the absorbance (optical density) of Calibration
Plate wells C01, D04, E02, F05, G03 and H06 with known standard values contained
in the Specification Sheet that accompanies each Calibration Test Plate.
•
Linearity: If the accuracy specifications are met, then the reader also proves to be
linear. Linearity can also be proven by performing a regression analysis on the OD
values in a program such as Microsoft® Excel. See the instruction on the following
page.
PowerWaveX Operator’s Manual
4-11
In Microsoft® Excel, open a spreadsheet and label one column “Assigned” and the
next column “Observed.”
1.
Enter the Assigned OD data for each glass filter in the first column from the
data sheet provided with the Test Plate.
2.
(Analyze one wavelength at a time.) Next, enter the Observed OD values for
the same glass filters in the adjacent column.
3.
Under Tools, select Data Analysis, and then Regression.
4.
Use the Regression “Input” box to enter the Assigned values as the “Input Y
Range” and the Observed OD as the “Input X Range.”
5.
Click the OK box and the Summary Output sheet will be displayed. An R
Square value of at least 0.99 is expected and the results will often be 0.999 or
better.
•
Repeatability: Repeatability is a measure of the instrument's ability to read the same
well with minimum variation between two reads with the well in the same location.
•
Turnaround: (Second Repeatability Result) This test confirms the reader's ability to
read samples accurately in different plate positions. Each channel is compared to at
least one other channel in this test: A to H, B to G, C to F, and D to E. This ensures
that all channels have comparable performance.
4-12
Performance Verification/Qualification Tests
Wavelength Accuracy
The C6 filter should be scanned between 580 and 590 nm in 1-nm increments using KC4 or
KCjunior software. If neither KC4 nor KCjunior is available, select six wavelengths at 1-nm
increments near the expected peak. The wavelength of the maximum absorbance should be
compared with the wavelength written on the sheet supplied with the test plate (see Figure 4-4
below). The accuracy of the wavelength should be ± 3 nm (± 2 nm instrument, ± 1 nm filter
allowance).
Figure 4-4: Sample data sheet showing wavelength of peak in the interval between
580 and 590 nm
For example:
If the data sheet value is 587 nm and the allowable range is ± 3 nm, and if the reader reports a peak
value of 590 nm, then the reader meets its specifications. If the reader reports 591 nm, then the
reader does not meet specifications.
PowerWaveX Operator’s Manual
4-13
Liquid Testing
Liquid testing tests the reader in ways that the Universal Test Plate cannot. The test plate will
indicate the absolute amount of light absorbed which will accurately test the linearity of the
electronics. The liquid test will help detect optical defects such as dirt or contamination that can
contribute to errant readings.
•
If you have the Universal Test Plate, you will need to run the simple Liquid Test 1 for
routine testing.
•
If you do not have a Universal Test Plate, test the linearity and repeatability of the
reader by preparing a series of solutions of varying absorbances as described in Liquid
Test 2.
•
If you prefer, you may use the described dye solution (see Table 4-3). The purpose of
the recipe is to create a solution that absorbs light in a well-defined manner and yields
~ 2.000 OD at full strength when dispensed at 200 microliters in a microplate well.
•
Alternatively, any solution that gives a stable color will suffice. (This includes
substrates incubated with an enzyme preparation and then stopped with an acidic or
basic solution.) Some enzyme/substrate combinations are shown below.
•
If you need to test a Universal Test Plate reader’s performance at 340 nm, run optional
Liquid Test 3 (see Table 4-5).
Table 4-2
Typical Enzyme-Substrate Combinations and Stopping Solutions
Enzyme
4-14
Substrate
Stopping Solution
Alkaline Phosphate
o-nitrophenyl phosphate
3N sodium hydroxide
beta-Galactosidase
o-nitrophenyl -beta-D
galactopyranoside
1M sodium carbonate
Peroxidase
2,2'-Azino di-ethylbenzothiazolinesulfonic acid (ABTS)
citrate-phosphate buffer, pH 2.8
Peroxidase
o-phenylenediamine
0.03N sulfuric acid
Performance Verification/Qualification Tests
STOCK SOLUTION FORMULATION ___________________________________________
The stock solution for Liquid Tests No. 1 and No. 2 may be formulated from the chemicals listed
below, or by diluting a dye solution available from Bio-Tek. See Procedure A or B outlined below
for details.
Procedure A
Required Materials:
•
Deionized water
•
FD&C Yellow No. 5 dye powder (typically 90% pure)
•
Tween® 20 (Polyoxyethylene sorbitan monolaurate, a wetting agent)
•
Analytical balance
•
1-liter volumetric flask
Table 4-3
Stock Solution Formulation for Liquid Test Nos. 1 and 2
FD&C Yellow No. 5 powder
0.092 g
Tween® 20
0.5 ml
DI Water to bring volume to:
1000 ml
Preparation of Stock Solution:
1.
Weigh out 0.092 gram of FD&C No. 5 yellow dye powder into a weigh boat.
2.
Rinse the contents into a 1-liter volumetric flask.
3.
Add 0.5 ml of Tween 20.
4.
Make up to 1 liter with DI water; cap and shake well.
This should create a solution with an absorbance of about 2.000 when using
200 µl in a flat-bottom microwell. The OD value result will be proportional to
the volume in the well and the amount of FD&C No. 5 dye used. You can use
a larger or smaller well volume, or add more dye or water to adjust the
solution. Note that too small a well volume may result in increased pipettingrelated errors.
PowerWaveX Operator’s Manual
4-15
Procedure B
Required Materials:
•
Bio-Tek QC Check Solution No. 1 (P/N 7120779, 25 ml; or 7120782, 125 ml)
•
Deionized water
•
5-ml Class A Volumetric Pipette
•
100-ml Volumetric Flask
Preparation of Stock Solution:
1.
Pipette a 5-ml aliquot of Bio-Tek QC Check Solution No. 1 into a 100-ml volumetric
flask.
2.
Make up to 100 ml with DI water; cap and shake well.
This should create a solution with an absorbance of about 2.000 when using
200 µl in a flat-bottom microwell. The OD value result will be proportional to
the volume in the well and the amount of QC Check Solution No. 1 used. You
can use a larger or smaller well volume, or add more Check Solution or water to
adjust the stock solution. Note that too small a well volume may result in
increased pipetting-related errors.
4-16
Performance Verification/Qualification Tests
LIQUID TEST 1 __________________________________________________________
This procedure will test for repeatability and consistency, making evident any problems with the
optics of the system.
1.
Using a freshly prepared stock solution (see Procedure A on page 4-15 or Procedure B
on page 4-16), prepare a 1:2 dilution using deionized water (one part stock, one part
deionized water; the resulting solution is a 1:2 dilution).
The concentrated stock solution will have an optical density of approximately
2.000 Abs. This value is not critical, but should be at the higher end of this
absorbance range. Should it exceed the range, simply reduce the volume in the
microwell. The diluted solution should have ODs of half of the concentrated
solution.
2.
Pipette 200 µl of the concentrated solution into the first column of wells of a new
96-well, flat-bottom microplate (Costar® #3590 is recommended). A new microplate is
required for this test, as any scratches may cause variations in the turnaround reading.
3.
Pipette 200 µl of the diluted solution into the second column of wells.
4.
Read the microplate five times at 405 nm using normal mode, single wavelength, no
blanking.
5.
Next, turn the microplate around so that well A1 is now in the H12 position and read
the plate five more times.
6.
The plate data can be exported to an Excel® spreadsheet using KC4 or KCjunior. The
mathematical computations described below may then be performed and the template
kept for future data reduction.
Channel-to-Channel Variation:
7.
Calculate the means of the wells in columns 1 and 2 in the normal plate position, and
in the turnaround position.
This test results in four comparisons of each channel to its corresponding channel, two
in column 1, and two in column 2.
Compare the mean reading for well A1 to its mean reading when in the H12 position.
Next, compare the mean values for the other wells to their corresponding mean values
with the well in the turnaround position. (Compare B1 to G12, C1 to F12, D1 to E12,
E1 to D12, F1 to C12, G1 to B12, H1 to A12, A2 to H11, B2 to G11, etc.). The
difference in the values for any two corresponding wells should be within the accuracy
specification for the instrument.
PowerWaveX Operator’s Manual
4-17
For example:
If the mean of well A1 in the normal position is 1.902, where the specified accuracy
is ± 1% ± 0.010 Abs, then the expected range for the mean of the same well in the
H12 position is 1.873 to 1.931 Abs. (1.902 * 1% = 0.019 + 0.010 = 0.029, which is
added and subtracted from 1.902 for the range.)
If any set of well values is out of the expected range, review the other three sets for
the same channel pair. Thus, if A1 and H12 are not within range of each other, review
the compliance of H1 to A12, A2 to H11, and H2 to A11. This will confirm that there
is a problem in one of the eight read channels or indicate that the result of one set of
wells was in error. If two or more sets of well values for a channel pair are out of the
allowed accuracy range, there may be contamination on, or a problem with, one of the
lenses.
Accuracy Specification:
For comparison in this test, the following accuracy specifications are applied,
using Normal mode and a 96-well microplate.
± 1% ± 0.010 Abs from 0.000 to 2.000 Abs
± 3% ± 0.010 Abs from 2.000 Abs to 3.000 Abs
4-18
Performance Verification/Qualification Tests
LIQUID TEST 2 __________________________________________________________
The recommended method of testing the instrument performance is to use the Universal Test Plate
to confirm alignment, repeatability, and accuracy, which will also confirm linearity.
If a Test Plate is not available, Liquid Test 2 can be utilized for these tests.
Preparation of Dilutions:
1.
Set up a rack containing 10 tubes, numbered consecutively.
2.
Prepare a concentrated stock test solution insert using either Procedure A on page
4-15, or Procedure B on page 4-16.
3.
Create a percentage dilution series, beginning with 100% of the original concentrated
stock solution in the first tube, 90% of the original solution in the second tube, 80% in
the third tube, all the way to 10% in the last tube. Use Class A volumetric pipettes for
better accuracy.
4.
Dilute using amounts of the remaining 0.05% solution of deionized water and
Tween® 20, as shown in Table 4-4.
Table 4-4
Test Tube Dilutions
Tube Number
1
2
3
4
5
6
Volume of Original
Solution (ml)
20
18
16
14
12
10
8
6
4
2
0
2
4
6
8
10
12
14
16
18
2.0
1.8
1.6
1.4
1.2
1.0
0.8
0.6
0.4
0.2
Volume of 0.05%
Tween Solution (ml)
Absorbance expected
if original solution is
2.0 at 200 µl
7
8
9
10
Plate Preparation:
5.
Pipette 200 µl of the concentrated solution from tube 1 into each well of the first
column, A1 to H1, of a new flat-bottom microplate (Costar® #3590 is recommended).
Next, pipette 200 µl from each of the remaining tubes into the wells of the
corresponding column of the microplate (tube 2 into wells A2 to H2, etc.).
Note:
The choice of dilutions and the absorbance of the original solution can be
varied. Use Table 4-4 as a model for calculating the expected absorbances of
a series of dilutions, given a different absorbance of the original solution.
PowerWaveX Operator’s Manual
4-19
Linearity – Test A
1.
Read the microplate prepared above using a normal mode dual wavelength at 450 nm
with 630 nm as the blank. Repeat the read four times for a total of five reads.
2.
The plate data can be exported to an Excel® spreadsheet using KC4 or KCjunior. The
mathematical computations described below may then be performed and the template
kept for future data reduction.
3.
Calculate the mean absorbance for each well, and average the means for each
concentration.
4.
Perform a regression analysis on the data to determine if there is adequate linearity.
For example:
In Microsoft Excel, under Tools, select Data Analysis and then Regression. (Prior to
opening the regression analysis tool, enter the expected results of the dilutions in a
row of cells to use in the analysis.) Use the Regression “Input” box to enter the
expected values as the “Input Y Range” and the mean absorbance for each
concentration as the “Input X Range.”
Expected results:
Since it is somewhat difficult to achieve high pipetting accuracy when conducting
linear dilutions, an R Square value of at least 0.99 is considered adequate.
Repeatability – Test B
1.
Calculate the mean and standard deviation for the five readings taken above at each
concentration. Only one row of data needs to be analyzed.
2.
For each mean below 2.000 Abs, calculate the allowed deviation using the
repeatability specification for a 96-well format of ±1% ± 0.005 Abs. If above 2.000
Abs, apply the ±3% specification.
3.
The standard deviation for each set of readings should be less than the allowed
deviation.
For example:
Absorbance readings of 1.950, 1.948, 1.955, 1.952, and 1.950 will result in a mean of
1.951, and a standard deviation of 0.0026. The mean (1.951) multiplied by 1%
(1.951 * 0.010) = 0.0195, which, when added to the 0.005 (0.0195 + 0.005) =
0.0245 Abs, which is the allowable deviation. Since the standard deviation is less
than this value, the reader meets the test criteria.
4-20
Performance Verification/Qualification Tests
Repeatability Specification:
±1% ± 0.005 Abs from 0.000 to 2.000 Abs
±3% ± 0.005 Abs from 2.000 Abs to 3.000 Abs
Channel-to-Channel Variation and Alignment – Test C
1.
Using the plate prepared for Test A above, conduct a turnaround test by reading the
plate with the A1 well in the H12 position five times. This test results in four
comparisons of each channel to its corresponding channel, two in column 1, and two
in column 2.
2.
Calculate the means of the wells in columns 1 and 2 in the normal plate position (data
is from Test A) and in the turnaround position (from Step 1 above). Compare the mean
reading for well A1 to its mean reading when in the H12 position. Next, compare the
mean values for the other wells to their corresponding mean values with the well in the
turnaround position. (Compare B1 to G12, C1 to F12, D1 to E12, E1 to D12, F1 to
C12, G1 to B12, H1 to A12, A2 to H11, and B2 to G11, etc.). The difference in the
values for any two corresponding wells should be within the accuracy specification for
the instrument.
For example:
If the mean of well A1 in the normal position is 1.902, where the specified accuracy
is ±1% ± 0.010 Abs, then the expected range for the mean of the same well in the
H12 position is 1.873 to 1.931 Abs. (1.902 * 1% = 0.019 + 0.010 = 0.029, which is
added and subtracted from 1.902 for the range.)
If any set of well values is out of the expected range, review the other three sets for
the same channel pair. Thus, if A1 and H12 are not within range of each other, review
the compliance of H1 to A12, A2 to H11, and H2 to A11. This will confirm that there
is a problem in one of the eight read channels, or indicate that the result of one set of
wells was in error. If any two sets of well values for a channel pair are out of the
allowed accuracy range, there may be contamination on, or a problem with, one of the
lenses.
3.
If the four corner wells are within the repeatability range, the reader is also in
alignment.
PowerWaveX Operator’s Manual
4-21
LIQUID TEST 3 __________________________________________________________
To verify operation of the PowerWaveX at 340 nm, perform the Liquid Test 3 Procedure described
on page 4-23. This test is optional as the front end of the instrument shows good correspondence at
various wavelengths. Thus, if the device performs properly using the test plate at 405 nm, it will
also perform adequately at 340 nm.
Required Materials:
•
Deionized Water
•
Pipettes
•
Costar® #3590 Flat-Bottom Microplate
•
Beakers and Graduated Cylinder
•
Analytical balance
•
Tween® 20 (Polyoxyethylene sorbitan monolaurate)
•
Phosphate Buffered Saline with Tween 20 (PBS Buffer Solution). Use
Sigma® P 3563 packets, which will be adequate for one liter of PBS solution each
(Procedure B) or prepare a 10X concentrate per Table 4-5 (Procedure A).
•
D-NADH Powder (D-Nicotinamide Adenine Dinucleotide, Reduced Form) Sigma®
bulk catalog number N 8129, or pre-weighed 10-mg vials, Sigma® number 340-110
Stock Buffer Solution Formulation:
Table 4-5
Phosphate Buffered Saline 10X Concentrate Solution
4-22
KH2PO4 anhydrous
0.2 grams
NaCl
8.0 grams
Na2HPO4 anhydrous
1.15 grams
KCl
0.2 grams
Tween® 20
0.5 ml
Add Deionized water to bring to
100 ml
Performance Verification/Qualification Tests
Procedure A:
Mix 45 ml of deionized water with 5 ml of the concentrated PBS solution (from Table 4-5)
in a beaker. Add 10 mg of the D-NADH powder and mix thoroughly. This is the high-level
test solution.
Procedure B:
Add 50 ml of a PBS solution (prepared from the Sigma powder) to a beaker. Add 10 mg of
the D-NADH powder and mix thoroughly. This is the alternate high-level test solution.
Liquid Test 3 Procedure:
1.
Check the absorbance of a sample of either high-level test solution created in
Procedure A or B at 340 nm on the microplate reader. This solution will have an
optical density (absorbance) of approximately 0.700 to 1.000. This value is not
critical, but it should be within this absorbance range. Adjust up by adding D-NADH
powder, if low, until the high-level test solution is at least at the lower end of this
range. Do not adjust if slightly high.
2.
Carefully prepare a mid-level test solution by diluting 15 ml of the high-level test
solution with 5 ml of the Sigma PBS solution. (If using the 10X-concentrate PBS
solution, you should mix one part of the concentrate with nine parts of deionized water
to obtain a low-level buffer similar to the Sigma PBS. Then use 5 ml of this solution as
the diluent.) This will be the mid-level solution.
3.
Carefully prepare a low-level test solution by diluting 10 ml of the high-level test
solution with 10 ml of the Sigma PBS solution. (If using the 10X-concentrate PBS
solution, you should mix one part of the concentrate with nine parts of deionized water
to obtain a low-level buffer similar to the Sigma PBS. Then, use 10 ml of this solution
as the diluent.) This will be the low-level solution.
4.
Pipette 150 µl of the concentrated solution into each well of the first two columns,
A1 to H1 and A2 to H2, of a flat-bottom microplate (Costar® #3590 is recommended).
Next, pipette 150 µl from the mid-level solution into the wells of columns 3 and 4 of
the microplate. Finally, pipette 150 µl of the low-level solution into the wells of
column 5 and 6 of the microplate.
5.
Read the microplate using Normal mode, single wavelength at 340 nm, no blanking
(or blank on air). Repeat the read four times for a total of five reads.
PowerWaveX Operator’s Manual
4-23
Repeatability – Test A
1.
The plate data can be exported to a Microsoft Excel spreadsheet using KC4™ or
KCjunior™. The mathematical computations described below may then be performed
and the template kept for future data reduction.
2.
Calculate the mean and standard deviation for the five readings of each well.
3.
For each mean, calculate the allowed deviation using the repeatability specification for
a 96-well format of ±1.0% ± 0.005 Abs.
4.
The standard deviation for each set of readings should be less than the allowed
deviation.
For example:
Absorbance readings of 0.802, 0.802, 0.799, 0.798, and 0.801 will result in a mean of
0.8004, and a standard deviation of 0.0018. The mean (0.8004) multiplied by 1%
(0.8004 * 0.010) = 0.008, which, when added to the 0.005 (0.008 + 0.005) = 0.013,
which is the allowable deviation. Since the standard deviation is less than this value,
the reader meets the test criteria.
Linearity – Test B
1.
Obtain an average mean for each concentration by averaging the mean values for each
well that were obtained above.
2.
Perform a regression analysis on the data to determine if there is adequate linearity.
For example:
In Excel, under Tools, select Data Analysis and then Regression. (Prior to opening the
regression analysis tool, enter the expected results of the dilutions in a row of cells to
use in the analysis.) Use the Regression “Input” box to enter the expected values as the
“Input Y Range” and the mean absorbance for each concentration as the “Input X
Range.”
Expected results:
Since it is somewhat difficult to achieve high pipetting accuracy when conducting
linear dilutions, an R Square value of at least 0.99 is considered adequate.
4-24
Performance Verification/Qualification Tests
Maintenance of Robotic Units
Use the following procedure for periodic lubrication of PowerWaveX robotic units.
1.
Remove the shroud.
2.
Lubricate the components shown in Figures 4-5 and 4-6 every six months, or after
10,000 cycles, according to the following instructions. Use Bio-Tek lubricant PN
66039.
Componen
t
Location
Procedure
A
Lubricate the underside of the bracket.
B
Lightly lubricate the shaft indicated.
Apply a heavy coating of lubricant to both sides of the motor shaft. Run the motor
shaft back and forth through the motor to ensure that the internal drive nut is
heavily lubricated. Use the robotic motor adjustment procedure provided below to
work the lubricant into the internal drive nut.
C
Ensure that the mylar sheet (BTI P/N 7262113) is in place on top of the main PCB
and below the motor assembly before lubricating the motor. Tape to the chassis
with black electrical tape. (This may already be in place.)
D
E
Lightly lubricate the surface of the roller.
F
Lightly lubricate the underside of the hook-in bracket.
3.
Reassemble the shroud.
4.
After lubrication/reassembly, the robotic door needs to be set to the open and closed
positions.
5.
Press the following keys to adjust the door. At the Main Menu:
Press the UTIL soft key.
Press the Setup soft key.
Press the hidden key between the Main Menu and Previous Screen keys.
6.
You should see the Door Adjustment screen.
7.
Press the CLOSE soft key.
PowerWaveX Operator’s Manual
4-25
8.
Press the UP soft key until you just see the door move up. You may have to press the
key many times before the door moves. When the door does move, press the DOWN
soft key 4 times. This positions the door lift ram just behind the door roller in the
closed position.
9.
Press the OPEN soft key.
10. Press the UP soft key to raise the door to the desired height. Press the DOWN soft key
if you want to adjust the door downward.
11. Press the CLOSE soft key. This will close the door. The door should rest closed on the
top shroud.
12. Press the Main Menu key.
4-26
Performance Verification/Qualification Tests
D
C
A
B
Figure 4-5: Location of bracket, motor shaft, mylar sheet, and main PCB/motor assembly
PowerWaveX Operator’s Manual
4-27
Figure 4-6: Roller surface and hook-in bracket
4-28
Performance Verification/Qualification Tests
Appendix A
Decontamination and Cleaning
This appendix contains the procedures for
decontaminating and cleaning the PowerWaveX.
Decontamination Procedure
If the PowerWaveX is to be shipped after being exposed to potentially hazardous material, it should
be decontaminated. The following procedure outlines how to decontaminate the instrument
before packaging and shipment.
PURPOSE ________________________________________________________________
Decontamination minimizes the risk to all who come in contact with the reader during shipping,
handling, and servicing. It is also required by the U.S. Department of Transportation regulations.
GENERAL CONSIDERATIONS _________________________________________________
•
Any laboratory instrument that has been used for clinical analysis is considered a
biohazard and should be decontaminated prior to handling. Intact skin is generally
considered an effective barrier against infectious organisms; however, small abrasions
and cuts may not be always be visible. Prophylactic gloves must be worn when
handling instruments that have not been decontaminated. Gloved hands should be
considered contaminated at all times and must be kept away from eyes, mouth and
nose at all times.
•
Mucous membranes are considered prime entry routes for infectious agents. Wear eye
protection and a surgical mask when there is a possibility of aerosols.
•
iii
Eating and drinking while decontaminating instruments is not advisable.
Important! Disconnect the unit from the power supply for all decontamination or
cleaning operations.
PROCEDURE _____________________________________________________________
Warning! The bleach solution is caustic; wear gloves and eye protection when handling the
solution. Do not soak the instrument keypad – this will cause damage. Wipe the keypad with a
damp cloth.
1.
Turn off and unplug the instrument.
2.
Prepare an aqueous solution of 0.5% Sodium Hypochlorite (NaClO, or bleach).
Be sure to check the % NaClO of the bleach you are using; this information is
printed on the side of the bottle. Commercial bleach is typically 10% NaClO;
if this is the case, use a 20:1 mixture. Household bleach is typically 5%
NaClO; if this is the case use a 10:1 mixture.
3.
Wipe down the carrier and all exposed surfaces of the unit with the bleach solution.
4.
Discard the used gloves and towels.
Cleaning Procedure
Exterior surfaces may be cleaned (not decontaminated ) with a cloth moistened (not soaked) with
water or water and mild detergent. Do not immerse instrument or spray with liquids.
A-2
Decontamination and Cleaning
Appendix B
Computer Control
The PowerWaveX can be controlled either from the
reader's front panel or from a computer connected to the
reader via the computer's serial port. This chapter
describes the features of computer control, and explains
how to configure the computer to control the reader.
Overview
With the computer control feature, the user is provided even more power and flexibility.
For example, the PowerWaveX can run computer-controlled kinetic assays using up to six
wavelengths on the same microplate. Spectral scan assays may be performed that use the entire
wavelength range from 200 nm to 999 nm, at 1 nm increments.
When using computer control, the blanking and data reduction capabilities on the PowerWaveX are
suppressed, and all data is returned to the user for evaluation. Readings higher than 3.000 OD may
be transmitted.
This appendix details the protocols necessary for communicating with the PowerWaveX reader. In
addition, instructions are provided for controlling the reader with Bio-Tek's KCjunior or KC4
software packages.
•
Controlling the Reader with KC4, page B-2.
•
Controlling the Reader with KCjunior, page B-4.
•
Controlling the Reader using Serial Protocol, page B-6.
•
Using the Stop Key to Halt Plate Scans and Reads, page B-20.
•
Status String Format, page B-21.
Controlling the Reader with KC4™
The PowerWaveX can be operated using a computer running Bio-Tek's KC4 software. Follow the
steps below:
1.
Power off the computer and the reader. Connect the appropriate serial cable
(PN 75053) between the two machines. See Table B-1 on the following page for a
pin-out description of the cable.
2.
Power up both machines.
3.
Install KC4 on the computer’s hard drive.
4.
Once installed, start KC4.
5.
Select System, Readers.
6.
Scroll through the list of Available Readers and select the appropriate PowerWaveX
reader model. Click the Port button (and subsequent Setup button), to define the
following communications parameters:
• Port:
COM1, 2, 3, or 4 (the serial port used for the
RS-232 cable connection).
7.
• Baud Rate:
9600
• Data Bits:
8
• Parity:
None
• Stop Bits:
2
Click the Current Reader button to attempt to establish communications with the
reader, using the currently defined communication parameters.
8.
If the test passes, click OK to save the settings and close the dialog box. If the test
fails, KC4 will provide appropriate instructions for resolving any problems. See also
the Problems section below.
To learn more about KC4 and how to define assays and read plates, refer to the KC4 User’s
Manual.
Problems
If KC4 fails to communicate with the reader and it displays a serial communications error, check the
cable plug-in location to ensure that it matches the setup choices and is not a Null cable. If this is
suspected, add another Null and try again.
If an Incorrect Reader Model Connected dialog box is displayed, click OK to clear the screen and
select System, Readers, Available Readers. Verify that the reader selected is correct.
B-2
Computer Control
Table B-1
Serial Cable Pin-Out Description
Serial Cable Pin-Out*
PC (9-pin female)
Reader (25-pin female)
1+6
2
3
5
7
8
Shell
4
2
3
7
5+6+8
20
Shell (shielding)
* For a 25-pin PC connection using Bio-Tek serial cable
(PN 75053) plus 9pM-25pF adapter (PN 49755).
Getting Started with KC4
The following instructions briefly describe how to read a plate using KC4. Refer to KC4's Help
system and User's Guide early and often to learn how to create protocols, assign well identifiers,
read plates, print reports, and more.
To read a plate using KC4:
1.
Select Data|New Plate.
2.
If prompted to select a protocol, select “Empty Protocol” and click OK. If not
prompted, select Protocol|New, or use KC4’s Protocol Wizard to step through protocol
creation.
3.
Select Protocol|Reading. The Reading parameters dialog will appear.
4.
Select a Reading Type of Endpoint, Kinetic, or Spectrum.
5.
Define the wavelength(s) at which the plate will be read.
6.
Select a Plate Geometry of 8x12 (96-well plate) or 16x24 (384-well plate).
7.
Define other reading parameters as necessary. Click the Help button for assistance.
8.
When complete, click OK.
9.
Select Data|Read Plate. The Plate Reading dialog will appear.
10. Enter any comments, place the plate on the carrier, then click Start Reading to begin
the plate read.
•
The plate will be read and then the raw data results will display in KC4.
•
To analyze, manipulate, or print results, protocol parameters should be
defined. Refer to KC4's Help system or User's Guide for instructions.
PowerWaveX Operator’s Manual
B-3
Controlling the Reader with KCjunior™
The PowerWaveX can be operated using a computer running Bio-Tek's KCjunior software. Follow
the steps below:
1.
Power off the computer and the reader. Connect the serial cable (PN 75053) between
the two machines. See Table B-1.
2.
Power up both machines.
3.
Install KCjunior on the computer’s hard drive.
4.
Once installed, start KCjunior.
5.
Select Setup, then Reader1. To select the PowerWaveX reader and define the
communications parameters, choose the following setup parameters:
• Reader:
PowerWaveX
• Com Port:
COM1 or COM2 (the serial port used for
the RS-232 cable connection)
• Baud Rate:
9600
• Data Bits:
8
• Parity:
None
• Stop Bits:
2
• EOT Character: Keep the default number.
6.
Click the Test Communications button to attempt to establish communications with
the reader, using the currently defined communication parameters. If a Serial Write
Error dialog box is displayed, an incorrect Com Port may have been selected. Select a
different port and then repeat this step.
7.
If the test passes, click OK to save the settings and close the dialog box. If the test
fails, follow the directions provided by KCjunior, then click Test Communications
again.
To learn more about KCjunior and how to define assays and read plates, refer to the KCjunior
User’s Manual.
Problems
If KCjunior fails to communicate with the reader, and displays a serial communications error, check
the cable plug-in location to make sure it matches the setup choices and is not a Null cable. If this is
suspected, add another Null and try again.
B-4
Computer Control
Getting Started with KCjunior
The following instructions briefly describe how to read a plate using KCjunior. Refer to KCjunior's
Help system and User's Guide early and often to learn how to create protocols, assign well
identifiers, read plates, print reports, and more.
To read a plate using KCjunior:
1.
Click Read Plate from KCjunior's main screen. The Read Plate Dialog will appear.
2.
If desired, enter a Results ID and a Plate Description, and then click Read Plate. The
Protocol Definition dialog will appear.
3.
Select a Read Method Type of Endpoint, Kinetic, Multi-Wavelength, or Spectrum.
4.
Define the wavelength(s) at which the plate will be read.
5.
Select a Plate Geometry of 8x12 (96-well plate) or 16x24 (384-well plate).
6.
Define other reading parameters as necessary. Click the Help button for assistance.
7.
When complete, click OK to return to the Read Plate dialog. If desired, enter a Plate
ID.
8.
Place the plate on the carrier, then click OK to start the plate read.
•
The plate will be read and then the raw data results will display in KCjunior.
Print the raw data by selecting Plate|Print Results.
•
To analyze or manipulate results, a protocol should be defined. Refer to
KCjunior's Help system or User's Guide for instructions.
PowerWaveX Operator’s Manual
B-5
Controlling the Reader Using Serial Protocol
At baud rates of 1200, 2400, and 9600, the PowerWaveX is capable of sending and receiving data
through its serial port (RS-232C). The baud rate used for transmission is held in nonvolatile
memory and can be changed by the user. Other serial port parameters, No Parity, 8 Data Bits, and 2
Stop Bits are fixed and cannot be changed.
The RS-232C serial port on the PowerWaveX is configured as a DTE (see the section Setting Up the
Serial Port for Communications in Chapter 2); that is, the unit is wired to look like a modem. Data
is received on Pin 3 (the RX Pin), and transmitted on Pin 2 (the TX pin).
Computer Control Command Set
A command from the computer to the reader consists of a single ASCII character, and in some
cases, subsequent argument data. Upon receipt of a valid command character, the reader returns an
<ACK> character. Some commands also return response data to the host computer. Upon
completion of command processing, the PowerWaveX transmits a status string to the computer.
While awaiting a command, the PowerWaveX responds to nulls or other unexpected characters by
clearing its input buffer and transmitting a <NAK>. Therefore, if valid commands are preceded by
invalid characters, they may be missed.
Refer to Table B-2 below for the ASCII Control Characters used in the computer control protocol.
Table B-2
ASCII Control Characters Used in Computer Control Protocol
B-6
ASCII
Code
Function
Hex
Code
Decimal
Code
Control
Code
Reader
<-- >
ACK
Acknowledge
06
06
^F
--->
NAK
Negative
acknowledge
15
21
^U
--->
RS
Record separator
1E
30
^^
--->
ETX
End of text
03
03
^C
<-->
DLE
Data link escape
10
16
^P
--->
CR
Carriage return
0D
13
^M
<---
LF
Line feed
0A
10
^J
<---
CTRL-Z
Control Z
1A
26
^Z
<---
Computer Control
All ASCII character strings representing numbers or names are transmitted most significant digit or
letter first. Data values not indicated as ASCII are treated as binary integers, and are transmitted
least significant byte first.
Lower wavelength limits shown are for the PowerWaveX 200 only. The PowerWaveX 340 is
limited to wavelengths above 340 nm, and will not accept lower values.
The following section describes the supported computer control command set.
STORE PLATE CARRIER (‘A’)
This command causes the plate carrier to move inside the instrument.
host:
valid limits:
response:
‘A’
no arguments
<ACK>
status string (5)
PRESENT PLATE CARRIER (‘J’)
This command causes the plate carrier to move back outside the instrument where it can be loaded
with a microplate.
host:
valid limits:
response:
‘J’
no arguments
<ACK>
status string (5)
PowerWaveX Operator’s Manual
B-7
CONFIGURE WAVELENGTHS (‘M’)
This command downloads the wavelength setup table for use in future read cycles.
host:
valid limits:
‘M’
response:
<ACK/NAK>
data bytes:
1-3
1st wavelength (“200” - “999”, ASCII)
4
comma separator (‘,’)
5 - 24
remaining wavelengths, commas
<ETX>
status string (5)
Each wavelength is represented by “xxx,” (3 ASCII digits, followed by a comma). Six wavelengths
must be sent (24 bytes), followed by ETX after the last comma. If a wavelength is not to be
specified, it must be replaced by “000”. The wavelength setup table is stored in non-volatile
memory, and only needs updating when a change in current wavelength configuration is desired.
After wavelength configuration, a self-test ('*' command) sequence must be run to calibrate gains
and generate self-check information for the new wavelength configuration. It is not absolutely
necessary to configure and calibrate wavelengths for use, but performance may be compromised
slightly if this is not done. The wavelengths stored are automatically calibrated during the
instrument power up self-test.
B-8
Computer Control
READ PLATE (‘S’)
This command causes a microplate to be read according to the currently loaded assay definition
table.
host:
valid limits:
response:
‘S’
no arguments
<ACK/NAK>
Reader response protocol (ASCII format):
status string (5)
for each wavelength specified (up to six):
wavelength start code (1)
<CR>
for each row:
for each column (well) within specified strip range:
comma separator (1)
','
sign (1)
'+' or '-'
data (4)
“1234”
row terminator (2)
<CR>, <LF>
data terminator (1)
<^Z>
checksum (1)
0 - 255 (not ASCII)
status string (5) (only if read error)
Absorbance data is returned in the form of a 5-character ASCII string, with a sign character
followed by four digits. A decimal point is not included, but should be inserted by the user after the
first digit, i.e. “+1234” should be translated as +1.234 OD. Overrange readings are indicated by the
5-character string “*****” instead of a signed numerical value.
Checksum calculation starts with the first byte AFTER the data start code, up through and including
the <^Z> data terminator code at the end of each wavelength’s plate data. The checksum is
transmitted as an integer data byte (not ASCII).
PowerWaveX Operator’s Manual
B-9
SET ASSAY DEFINITION (‘V’)
This command downloads to the reader a 170-byte assay definition table to be used in subsequent
plate reads.
Note: To minimize confusion, all “don’t cares” should be set to 0.
host:
valid limits:
‘V’
response:
<ACK/NAK>
data bytes:
1
don’t care
2-7
assay name (ASCII characters)
8 - 30
don’t care
31
encoded byte:
bits 0 - 3 (0x0F)
0
bit 4 (0x10)
1: shake, 0: no shake
bit 5 (0x20)
1: shake before every read, 0: before first read only
bit 6 (0x40)
1: continuous shake, 0: timed shake
bit 7 (0x80)
1: use total kinetic time, 0: use kinetic read count
32
number of scans per well for well scanning
(must be an odd integer from 1 to 31)
33 - 49
don’t care
50 - 55
“000000” (ASCII)
56
read type (0: endpoint, 2: kinetic, 3: scan)
Note: 3 indicates well scanning. To perform
spectral scanning, use the '&' SCAN PLATE
command.
B-10
57 - 59
don’t care
60 - 61
kinetic interval (seconds, 0 - 9999)
62 - 63
kinetic read count (2 - 9999)
64 - 66
don’t care
67 - 69
first wavelength (“200” - “999”, ASCII)
Computer Control
host:
valid limits:
response:
70 - 84
remaining wavelengths (“000” - “999”, ASCII)
85 - 162
don’t care
163 - 164
shake time (seconds, 0 - 999)
165
shake speed (0: slow, 1: med, 2: fast, 3: variable)
166 - 167
total kinetic read time (minutes, 1 - 9999)
168
delay before first read (0: no, 1: yes)
169 - 170
delay (seconds, 0 - 999)
status string (5)
Up to six wavelengths may be specified for a single plate read definition.
The assay definition table must be downloaded before other plate-specific commands are sent, such
as “Set Plate Geometry” and “Set Strip Range”. If a wavelength is not to be used, it must be
replaced by “000”.
GET WAVELENGTH TABLE (‘W’)
This command uploads the wavelength setup table from the reader.
host:
valid limits:
response:
‘W’
no arguments
<ACK/NAK>
Reader response protocol (ASCII format):
for each wavelength configured (six):
wavelength
(“000” - “999”)
comma terminator
‘,’
status string (5)
PowerWaveX Operator’s Manual
B-11
HALT (‘X’)
This command causes any read or spectral scan in progress to be halted.
host:
valid limits:
response:
‘X’
no arguments
none (see below)
no status string
The following events occur when this command is invoked:
1.
The scan or read process is halted, and all axes are returned to their home positions (the plate is
be moved back outside where it can be accessed).
2.
The reader transmits a <DLE> character to the computer when the above process is completed.
3.
No more data is transmitted by the read or scan in progress, nor is a status string.
SET PLATE GEOMETRY (‘{’)
This command selects a microplate geometry to be used with the currently loaded assay definition.
host:
valid limits:
‘{’
response:
<ACK/NAK>
data bytes:
1
geometry (2 or 3)
2: 96 wells (8 x 12)
3: 384 wells (16 x 24)
If no Set Plate Geometry command is sent after the assay definition table, the default 96 well plate
is assumed (it is recommended, however, that you issue this command even if using the 96 well
plate).
If only a range of strips is desired, the Set Strip Range command must be sent AFTER the Set Plate
Geometry command.
B-12
Computer Control
SET STRIP RANGE (‘%’)
This command selects a range of adjacent strips (numbered plate columns) to be read.
host:
valid limits:
‘%’
response:
<ACK/NAK>
data bytes:
1
first strip (1 - 24)
1
last strip (1 - 24)
status string (5)
By default the instrument is initialized to perform full plate reads, and automatically reverts to this
default when a new assay definition table is loaded, either from the front panel or by computer
control. The strip range is also reset when the Set Geometry command is sent.
GET MINIMUM KINETIC INTERVAL (‘$’)
This command provides a means for the user to determine the minimum kinetic read interval, as
dictated by the current assay definition table, strip range, and even baud rate selected.
host:
valid limits
response:
'$'
no arguments
<ACK/NAK>
kinetic interval (2)
status string (5)
The reader returns the minimum kinetic interval as a 2-byte integer value (not ASCII, so low byte
first).
PowerWaveX Operator’s Manual
B-13
SCAN PLATE (‘&’)
This command causes the reader to perform a spectral scan on the indicated microwell.
host:
valid limits:
'&'
response:
<ACK/NAK>
data bytes:
1-2
row (“00” - “16”, ASCII)
“00” indicates scan entire strip (column)
Note: Do not use 00 if reading 384-well plate.
3-4
column (“01” - “24”, ASCII)
5-7
start wave (“200” - “999”, ASCII)
8 - 10
stop wave (“201” - “999”, ASCII)
11 - 13
wave step (“001” - “799”, ASCII)
14
series option (‘0’ - ‘3’, ASCII)
'0': single scan
'1': first scan in series
'2': next scan
'3': last scan
15
calibrate option (‘0’ - ‘1’, ASCII)
'0': calibrate only if necessary
'1': calibrate before scanning
16 - 17
shake time in seconds (“00” - “99”, ASCII)
18
shake speed
'0': slow
'1': medium
'2': fast
'3': cycle through speeds listed above
<ETX>
B-14
<ACK/NAK>
Computer Control
host:
valid limits:
response:
Reader response protocol (ASCII format):
status string (5)
for each wavelength undergoing calibration (none to all waves selected):
wavelength (3)
“200” - “999”
wave terminator (1)
<CR>
calibration terminator (1)
<^Z>
for each wavelength specified for first pass scan (none, or all waves selected):
wavelength (3)
“200” - “999”
wave terminator (1)
<CR>
first pass terminator (1)
<^Z>
data start code (1)
<CR>
for each wavelength specified for scan:
wavelength (3)
“200” - “999”
for each well selected in strip (1 or 8 wells):
comma separator (1)
','
sign (1)
'+' or '-'
data (4)
“1234”
well/strip terminator (2)
data terminator
(1)
checksum (1)
<CR>, <LF>
<^Z>
0 - 255 (not ASCII)
status string (5) (only if error)
The reader sequentially scans the well or strip from the start wavelength through the stop
wavelength in increments dictated by the wavelength step. At least two wavelengths must be
scanned. The start must be less than the stop, and the step must be less than or equal to the
difference.
PowerWaveX Operator’s Manual
B-15
If the calibrate option is selected (‘1’), all wavelengths indicated for the current spectral scan will be
calibrated. If calibration is not selected, any wavelengths selected but not scanned since the
instrument was powered up will be calibrated anyway. In addition, any selected wavelengths with
previous errors detected will also be calibrated. If no calibration is performed at all, the <^Z>
terminator is returned alone with no wavelengths. Calibration is not absolutely necessary with each
plate, but is recommended in scans where absolute accuracy is a requirement.
Due to the fact that even-row detectors are offset left by one column, the reader uses two passes to
collect data when an entire strip is selected to be scanned. During the first pass, only the
wavelength being scanned is returned. Absorbance data is returned for each wavelength for the
entire strip during the second pass. The first pass terminator is always returned even if no first pass
is necessary.
If more than one well or strip is to be scanned, the series option can be used to make the process
more efficient. If a sequence of strips is to be scanned using the series option, they must be
contiguous and selected in sequence from left to right, since first pass data for the next strip is
collected during the second pass for the currently-selected strip. The last strip selected (last scan)
then requires no first pass. Calibration is only performed if the “single” or “first” scan option is
selected, and remains valid through the remaining wells or strips in the series.
Absorbance data is returned in the form of a 5-character ASCII string, with a sign character
followed by four digits. A decimal point is not included, but should be inserted by the user after the
first digit, i.e. “+1234” should be translated as +1.234 OD. Over-range readings are indicated by
the 5-character string “*****” instead of a signed numerical value.
Checksum calculation starts with the first byte AFTER the data start code, up through and including
the <^Z> data terminator code. The checksum is transmitted as an integer data byte (not ASCII).
If any invalid arguments are sent by the host computer, a <NAK> response is returned by the
reader.
The Set Geometry command must be sent before any Scan Plate commands if a microplate other
than the standard 96-well plate is to be used.
B-16
Computer Control
SELF-TEST (‘*’)
This command causes the reader to perform a system self-test and calibration. This should be
performed any time the wavelength configuration is changed (‘M’ command).
host:
valid limits:
response:
'*'
no arguments
<ACK/NAK>
data stream
status string (5)
The reader responds by sending a variable-sized stream of ASCII character data representing
various calibration and test results. This data stream is followed by the standard status response
string.
SET TEMPERATURE (‘[’)
This command sets the incubation chamber temperature setpoint.
host:
valid limits:
'['
response:
<ACK/NAK>
data bytes:
1-2
temperature setpoint (0, 22 - 50)
status string (5)
If the instrument is not equipped with a working incubator, or if the indicated temperature setpoint
is out of range, an incubator setpoint error will be returned with the standard status response string.
A setpoint of zero will turn the incubator heaters off.
PowerWaveX Operator’s Manual
B-17
GET TEMPERATURE (‘]’)
This command returns the current temperature in the incubation chamber.
host:
valid limits
response:
']'
no arguments
<ACK/NAK>
temperature (2)
status string (5)
The reader returns the current temperature as a 2-byte integer value (not ASCII, so low byte first).
Temperature is returned scaled up by 10, i.e. 370 indicates a temperature of 37.0 degrees Celsius. If
the instrument does not have incubation, or an incubation error has been detected, 0x0000 will be
returned as temperature data. An incubator temperature error will then be flagged and returned with
the standard status response string. Otherwise, the current temperature (averaged over four thermal
zones) is returned as defined above.
The reader will accept and process the Get Temperature command at any time, including during a
read cycle. The temperature response will NOT interrupt a plate data response stream, however. If
a Get Temperature command is received during a data transmission, it will not be processed until
the transmission has completed, i.e. after the <^Z> checksum combination has been sent.
GET PLATE BARCODE (‘s’)
This command returns the barcode for the microplate currently in the instrument carrier.
host:
valid limits
response:
's'
no arguments
<ACK/NAK>
barcode (33)
status string (5)
B-18
Computer Control
The barcode is returned as a 33-character string, with a null terminator (0x00) as the last character.
If the barcode is less than 32 characters it is null-terminated, then padded with blank characters
(0x20) up to the final null. If the barcode is more than 32 characters it is truncated.
If no barcode is detected after two passes of the microplate carrier in front of the scanner, the nullterminated string “NR” is returned, followed by the remainder of the 33-character string of blanks
and the final null. A barcode error is flagged and returned with the standard status response string.
GET INSTRUMENT CONFIGURATION (‘}’)
This command returns a 16-bit binary-encoded word defining the instrument configuration.
host:
valid limits
response:
'}'
no arguments
<ACK/NAK>
configuration (2)
no status string
Instrument configuration data is returned low byte first.
Encoding is defined as follows:
code:
description:
0x0030
PowerWave 200
0x0031
PowerWave 340
0x0032
PowerWavex
0x0038
PowerWave 200 w/ Incubator
0x0039
PowerWave 340 w/ Incubator
0x003A
PowerWavex w/ Incubator
0x003B
PowerWavex 340 w/ Incubator
0x0072
PowerWaveX Select
0x007A
PowerWaveX Select w/ Incubator
0x00B0
PowerWave 200 w/ Robot Access Door
PowerWaveX Operator’s Manual
B-19
code:
description:
0x00B1
PowerWave 340 w/ Robot Access Door
0x00B2
PowerWavex w/ Robot Access Door
0x00B8
PowerWave 200 w/ Incubator and Robotic Access Door
0x00B9
PowerWave 340 w/ Incubator and Robotic Access Door
0x00BA
PowerWavex w/ Incubator and Robotic Access Door
0x00F2
PowerWaveX Select w/ Robot Access Door
0x00FA
PowerWaveX Select w/ Incubator and Robotic Access Door
OPEN PLATE ACCESS DOOR (‘(‘)
This command opens the robotic plate access door
host:
valid limits
response:
'('
no arguments
<ACK/NAK>
status string (5)
CLOSE PLATE ACCESS DOOR (‘)’)
This command closes the robotic plate access door
host:
valid limits
response:
')'
no arguments
<ACK/NAK>
status string (5)
Using the Stop Key to Halt Plate Scans and Reads
Pressing the STOP key on the reader while a computer-control-initiated plate read or spectral scan
is in progress causes the following to occur.
1.
The scan or read process is halted, and all axes are returned to their home positions (the plate is
moved back outside where it can be accessed).
B-20
2.
The reader transmits a <DLE> character to the computer when the above process is completed.
3.
No more data is transmitted by the read or scan in progress, nor is a status string.
Computer Control
Status String Format
Following execution of each command, the PowerWave sends a status string back to the computer.
This string consists of 5 successive ASCII characters -- RS, S1, S2, S3, and ETX:
⇒ RS
A record separator that marks the beginning of the status string.
⇒ S1
Always ASCII zero (‘0’)
⇒ S2
A single digit, used as a reader fault or error code number.
• ERROR CODES
Error codes indicate the following:
‘0’: no fault or error
‘8’: instrument failure - perform self-test
‘9’: error in assay, scan, or table definition
‘A’: error in strip range selection
‘B’: incubator setpoint error
‘C’: incubator temperature error
‘E’: barcode error
⇒ S3
Always ASCII zero (‘0’)
⇒ ETX
End Of Text -- marks the end of the status block.
Instrument failures are usually accompanied by an error code on the reader display.
ELx Status Mode Format:
This status string consists of 5 successive ASCII characters – a four-byte string representing a
hexadecimal status code, and then ETX.
Items described in angle brackets (<>) are indicated by an ASCII digit replacing the last ‘0’
character in the status code.
Items described in curly braces ({}) are indicated by an ASCII digit replacing the next-to-last ‘0’
character in the status code.
Fatal errors indicate a hardware failure, also shown on the instrument display screen, and require
recycling of instrument power.
iii
Note: Errors listed on the following pages are common to all reader instruments, and may
not all be applicable to any single given reader.
PowerWaveX Operator’s Manual
B-21
Fatal Errors
TCB NOT AVAIL ERR
“A100”
// task control block not available
READ NOT AVAIL ERR
“A200”
// read already in progress
NOT AVAIL ERR
“A300”
// <device> not available
CHECKSUM ERR
“A400”
// failed code checksum test on powerup
DR ALLOC ERR
“A500”
// DR steps alloc/free error <assay num)
DFLASH TIMEOUT ERR
“A600”
// data flash write timed out
DFLASH ERR
“A700”
// data flash readback didn’t match write {test}<chip>
CFLASH TIMEOUT ERR
“A800”
// code flash write timed out
HEAP CORRUPTION ERR
“A900”
// memory allocation heap corrupted
ATOD ERR
“AA00”
// <device> A/D converter never saw ready transition
NO ERR
“0000”
// no errors detected
ABORT ERR
“0100”
// read function aborted
NO SENSOR ERR
“0200”
// <motor> didn't find opto-sensor transition
NO BEAM ERR
“0300”
// <motor> didn't find saturation transition
Non-Fatal Errors
B-22
MOTOR VERIFY ERR
“0400”
// <motor> failed positional verify
SATURATION ERR
“0500”
// A/D signal saturated <test type>
FILTER GAIN ERR
“0600”
// <filter> gain out of range
NOISE TEST ERR
“0700”
// reader {channel} failed noise test
OFFSET TEST ERR
“0800”
// reader {channel} failed offset test
DARK RANGE ERR
“0900”
// read-time {channel}<filter> dark out of range
AIR RANGE ERR
“0A00”
// read-time {channel}<filter> air/blank out
ASSAY NUM ERR
“0B00”
// invalid <assay number>
PRINT TIMEOUT ERR
“0C00”
// printer timed out
CAL CHECKSUM ERR
“0D00”
// failed calibration checksum test
WAVE NOT FOUND ERR
“0E00”
// wavelength not found in table <read filter>
FILTER SIGNAL ERR
“0F00”
// {channel}<filter> signal out of range
CNFG DATA ERR
“1000”
// necessary configuration data missing
CNFG CHECKSUM ERR
“1100”
// failed configuration checksum test
CAL DATA ERR
“1200”
// necessary calibration data missing
MOTOR NOT HOMED ERR
“1300”
// <motor> not homed successfully
INCUBATOR FAILURE
“1500”
// incubator failure {error code}<zone(s)>
SC ASSAY DEF ERR
“1600”
// computer control assay definition error
KIN INTERVAL ERR
“1700”
// interval too short for selected options
KIN COUNT ERR
“1800”
// too many kinetic intervals
MALLOC ERR
“1900”
// malloc failed
STORE CURVE ERR“1A00”
// store curve failure
GET CURVE ERR
“1B00”
// get curve failure
ATOD INIT ERR
“1C00”
// A/D calib STBY transition not detected
Computer Control
Non-Fatal Errors (continued)
RESULTS DATA ERR
“1D00”
// results data error
CLOCK ERR
“1E00”
// error in clock communications
OVERLAP ERR
“1F00”
// bandpass overlap in filterset
BARCODE ERR
“2000”
// no valid barcode detected
INVALID PARAM ERR
“2100”
// invalid parameter value selected
PMT ERR
“2200”
// PMT test signal too high <test type>
LAMP ERR
“2300”
// lamp control failure <test type>
SENSOR POS ERR
“2400”
// test sensor position incorrect <motor>
FLASH MISS ERR
“2500”
// motor went by flash location too soon
XY LIMIT ERR
“2600”
// physical limit exceeded for area scan request
PANEL METHOD ERR
“2700”
// <assay> method doesn’t match first panel assay
MOTOR TIMER ERR
“2800”
// <motor> timer not available
VREF ERR
“2900”
// voltage reference failed <test type>
PLATE JAM ERR
“2A00”
// <motor> didn’t find middle sensor
Test Type Codes (lowest digit in returned error code)
FAIL POWER 5V
‘1’
// 5V power failed
FAIL POWER 24V
‘2’
// 24V power failed
Motor Codes (lowest digit in returned error code)
Carrier X Axis
‘0’
Filter Wheel
‘1’
Robotic Door
‘2’
Monochromator
‘3’
Carrier Y Axis
‘4’
(‘R’ instruments only)
(Select only, ‘2’ on HT)
Incubator Codes (second lowest digit in returned error code)
Range Error
‘0’
Thermistor Error
‘1’
A/D Error
‘2’
Affected zones are encoded in the lowest digit returned – one bit per zone.
Data Flash Codes (2nd lowest digit in returned error code)
Readback Error
‘0’
// data read back didn’t match data written
Copy Error
‘1’
// final data readback didn’t match original passed in
A/D Device Codes (lowest digit in returned error code)
Absorbance measurement
‘1’
Fluorescence measurement
‘2’
Incubation measurement
‘3’
Voltage reference
‘4’
PowerWaveX Operator’s Manual
B-23
B-24
Computer Control
Appendix C
Error Codes
This chapter describes Error Codes that may appear on the reader.
If an error is displayed and you cannot solve the problem,
call Bio-Tek’s Technical Assistance Center. Refer to
Chapter 1, Technical Support for contact information.
Error Codes
An error code is displayed on the microplate reader as a four-digit identifier. The first digit is
usually 0 or A. A 0 denotes a non-critical error, which means the instrument will still respond to
keypad input. An A denotes a serious error, which requires that the reader be powered down, and
then powered back up before any diagnostics can be performed.
Displayed Error
Potential Cause
ERROR 0200
Plate carrier did not find the home sensor
ERROR 0201
Order sorting filter wheel did not find the home
sensor
ERROR 0202
Robot lid lift did not find the home sensor
ERROR 0204
Y axis could not find the optosensor
Errors 0200, 0201, 0202, and 0204 indicate that an axis was not able to correctly travel to
its “home” position. Both axes have optical sensors which, when interrupted, indicate that
the specific axis has been successfully homed.
PROBABLE CAUSE:
•
Carrier Axis Case - defective sensor. The X-axis movement is limited so that the
optical sensor cannot be interrupted.
•
Order Sorting Wheel Case - defective sensor. The order sorting filter wheel
movement is limited so that the sensor cannot be interrupted.
iii
Note: In cases where a sensor is not functioning, the motor will drive the
axis to its mechanical stop and generate substantial noise.
Displayed Error
Potential Cause
ERROR 0300
Carrier failed to find light beam
ERROR 0301
Order sorting filter wheel did not find home
ERROR 0303
Monochromator did not find home
Errors 0300, 0301, and 0303 indicate that a particular axis has moved to a point where the
light beam from the optics is no longer detectable by the measurement electronics.
PROBABLE CAUSE:
•
Carrier - A loose belt, loose motor pulley, or defective motor drive may cause the
carrier to ignore movement instructions.
•
Order Sorting Filter Wheel Motor - The filter wheel drive gear is loose or motor drive
failure is impeding filter wheel movement.
•
Flash Lamp – The flash lamp is not flashing due to a defective lamp, poor alignment,
or blocked optics.
Displayed Error
Potential Cause
ERROR 0400
Track carrier failed position verify
ERROR 0401
Order sorting filter wheel failed position verify
ERROR 0402
Robot lid lift failed position verify
ERROR 0403
Wavelength select (monochromator) failed
position verify
ERROR 0404
Y axis failed position verify (PowerWaveX Select
models only)
Errors 0400, 0401, 0402, 0403, and 0404 indicate that an axis failed its position verify test.
The position verify test verifies that the axis returns to the opto or the zero order (mono
case) in the same number of steps as it is expected to be away from home. If the axis does
not return home in the required number of steps, the test fails. Each axis is allowed to gain
or lose three steps.
PROBABLE CAUSE:
C-2
•
Belt slipping caused by incorrect tension, loose motor pulley, or loose belt clamp.
•
Defective motor drive circuit or Main PCB.
Error Codes
•
In the wavelength select case, failure to verify home correctly usually indicates an
alignment problem.
•
Anti-backlash spring on the monochromator has become free of the mounting post.
•
A defective flash lamp.
Displayed Error
Potential Cause
ERROR 0500
Saturation error
This error indicates that the optic system detected saturation conditions at the wavelength
selected during the read.
PROBABLE CAUSE:
•
Optic system is defective.
Displayed Error
Potential Cause
ERROR 0601
LAMBDA #1 Gain out of range
ERROR 0602
LAMBDA #2 Gain out of range
ERROR 0603
LAMBDA #3 Gain out of range
ERROR 0604
LAMBDA #4 Gain out of range
ERROR 0605
LAMBDA #5 Gain out of range
ERROR 0606
LAMBDA #6 Gain out of range
Errors 0601-0606 indicate that the gain for a specific wavelength is out of the range
necessary to ensure the filter’s performance to specifications.
PROBABLE CAUSE:
•
A defective flash lamp, misaligned optics, or defective analog board.
•
A defective order sorting filter.
PowerWaveX Operator’s Manual
C-3
Displayed Error
Potential Cause
ERROR 0700
Reader failed noise test
The last digit indicates the channel, 0-8, where 0 is the reference channel.
The reader noise test checks the DARK current signal level for stability. Dark current is
measured with the light blocked at maximum measurement channel gain. Four groups of
96 readings are taken at 100 ms intervals. This data is reduced to four averages that cannot
vary by 20 counts or a 0700 error will result. The outgoing production specification for
this test is 12 counts of variation. The unit will not flag an error until a 20-count variation
is seen.
PROBABLE CAUSE:
•
External signals getting into the measurement circuit. The bottom and top shrouds
should be correctly installed.
•
This problem in a correctly assembled unit could indicate a bad main or analog board.
Failure indicates excessive variation in the dark current (background) noise levels of
the measurement circuit.
•
Defective ground on the track zone 3 ground wire.
•
Malfunctioning incubation board.
•
Defective photodetector on the analog PCB.
Displayed Error
Potential Cause
ERROR 0800
Reader failed offset test
Error 0800 indicates that the measurement electronics’ dark current offset is outside of
acceptable limits at maximum gain. The noise signal level must be between 144 and 2019
counts.
PROBABLE CAUSE:
C-4
•
Ambient light leak (track access door open during self check).
•
The analog board could be dirty or defective.
•
The digital board power supplies could be outside their intended values.
•
The photodiode could be defective.
Error Codes
Displayed Error
Potential Cause
ERROR 0900
Read time dark value out of range
Error 0900 indicates that the dark current value taken during the current read is
significantly different from the same reading taken during the power-up self-check. The
last digit indicates the wavelength and the second to last digit indicates the channel.
PROBABLE CAUSE:
•
The measurement electronics background noise has changed since the last power-up
self-check. This could be caused by a large increase in external ambient light since
power-up. See Probable Cause for Error 0800.
•
Running the System Test will reset the value to the new baseline.
Displayed Error
Potential Cause
ERROR 0A00
Read time air blank out of range
Error A00 indicates that the blank (full signal) reading taken during the current read has
changed significantly from the same reading taken during the power-up self-check.
PROBABLE CAUSE:
•
The measurement electronics full signal level has changed since the power-up selfcheck was last run. The flash lamp could be near failure.
•
Running the System Test will restore to new baseline.
Displayed Error
Potential Cause
ERROR 0B00
Invalid assay
This error indicates that an assay number that is not programmed was selected.
PowerWaveX Operator’s Manual
C-5
Displayed Error
Potential Cause
ERROR 0C00
Printer timeout
Error 0C00 indicates that the printer in use is not responding.
PROBABLE CAUSE:
•
Printer not connected or powered up.
•
Printer’s parallel port may not be correctly selected in the on-board software. See
Printing and Data Communications in Chapter 2.
Displayed Error
Potential Cause
ERROR 0D00
Calibration checksum error
This error indicates that the stored checksum value for the calibration data does not match
the actual checksum. This indicates that the data in the quick flash memory has been
corrupted, probably due to an electronic defect.
Displayed Error
Potential Cause
ERROR 0E00
Wavelength not detected in reader’s Lambda table
Error 0E00 indicates that the specified assay wavelength is not in the Lambda table and
must be programmed in.
Displayed Error
Potential Cause
ERROR 0F00
Wavelength signal is out of specified range
This error indicates that the measurement frequency selected has a signal out of range. The
causes could be many. Essentially, the signal produced at a specific frequency is either too
high or too low to allow performance within the specification. The lamp could have gone
bad or a part of the optics system could be out of alignment. This can also be a flag for
filters starting to degrade.
C-6
Error Codes
Displayed Error
Potential Cause
ERROR 1000
Configuration data is missing
This error indicates that necessary configuration data is missing from memory, which
probably means it was never downloaded or it was downloaded incorrectly.
Displayed Error
Potential Cause
ERROR 1100
Failed configuration checksum test
This error indicates that the stored checksum value from the configuration data does not
match the actual checksum of the current configuration data. This means that the
configuration data has changed and the checksum stored is no longer valid. The error is
produced when outdated versions (old) of Extensions or Define Assay are used to create an
assay configuration file. This file is incompatible with the operation code within the
instrument’s memory. The fix for this problem is to recreate the assay definition on the
correct version of assay definition software and re-download it. This can also be caused by
a bad Flash memory. Reloading the basecode and assay configuration may fix this.
Displayed Error
Potential Cause
ERROR 1200
Calibration data missing
This error means that AUTOCAL has not been performed after a memory erase or in the
case of a new unprogrammed board immediately after the basecode or assay definition
download. The system must have the AUTOCAL sequence performed. This can also
indicate a bad Flash memory.
Displayed Error
Potential Cause
ERROR 1300
Motor not correctly homed
This error will occur if the error 0200 or error 0300 is ignored. The situation needs to be
fixed before the instrument is used.
PowerWaveX Operator’s Manual
C-7
Displayed Error
Potential Cause
ERROR 1400
Assay incubation error
Assay requires incubation but incubation is not available.
Displayed Error
Potential Cause
ERROR 1500
Assay incubation error
This error indicates that incubator failed to hold temperature within tolerances during the
assay.
Incubator Failure Error Data
Bits
Zone
Error
7
3
Temperature out of range
6
2
Temperature out of range
5
1
Temperature out of range
4
0
Temperature out of range
3
3
Thermistor out of range or shorted
2
2
Thermistor out of range or shorted
1
1
Thermistor out of range or shorted
0
0
Thermistor out of range or shorted
Examples:
Bit 6
Bit 0
↓
↓
Error code (1540) → 40 hex = 0100 0000
Zone 2 temp out of range, unknown reason (A/D failure?)
Error code (1544) → 44 hex = 0100 0100
Zone 2 temp out of range, due to zone 2 thermistor bad.
Error code (15FF) → FF hex = 1111 1111
All zones temp out of range, due to all thermistors bad.
Note: Any failure indicates a need to run a System Test and review of the printout (or KC4
screen).
C-8
Error Codes
Displayed Error
Potential Cause
ERROR 1600
Computer control assay definition error
Assay programming via computer control has tried to define an invalid assay sequence or
parameter.
Displayed Error
Potential Cause
ERROR 1700
Kinetic interval too short for selected options
ERROR 1800
Too many kinetic intervals programmed
ERROR 1900
Memory allocation failure
ERROR 1A00
Store curve error
ERROR 1B00
Get curve error
ERROR A100
Task control block error
ERROR A200
Reader function already in use
ERROR A300
Device not available
ERROR A400
Failed code checksum on power-up
ERROR A500
Power good error, power dropped below safe level
ERROR A600
Quick flash configuration time-out
PowerWaveX Operator’s Manual
C-9
C-10
Error Codes
Appendix D
Sample Reports
This appendix contains samples of the different
types of reports available on the PowerWaveX.
Reports are automatically generated after a plate has been read (see Specifying Data Output and
Reporting Options in Chapter 2 for information on selecting report formats).
In addition, Map, Assay, and Assay List reports can be printed via the REPORT option from the
Main Menu.
This Appendix contains samples of the different types of reports available on the PowerWaveX.
Figure D-1: Matrix Report showing well IDs, calculated OD values
and concentrations, calls on samples, and comments
D-2
Sample Reports
Figure D-2: Curve Fit Report showing the curve method, equation, coefficients, and r-squared value
PowerWaveX Operator’s Manual
D-3
Figure D-3: First page of Column Report, showing plate data for blanks and
Standards, the Interpretation of Results, and comments
D-4
Sample Reports
Figure D-4: Second page of Column Report, showing plate data for samples
PowerWaveX Operator’s Manual
D-5
Figure D-5: Column Report without Samples
.
D-6
Sample Reports
Figure D-6: Panel Report
PowerWaveX Operator’s Manual
D-7
Figure D-7: Assay Detail Report
D-8
Sample Reports
Assay List
Version: 2.0
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Raw_OD
Raw_OD w/blk
Bio-Cell Read
K-Factor Bio-Cell
DNA Quant
260/280 Ratio
Protein Quant
Oligo Quant
Protein Quant
Phenol
EDTA test
A320 Scattering
Lowry Protein
Bradford Protein
BCA Protein
B=galactosidase
NADH
HRP 5AS
HRP ABTS
HRP OPD-S
HRP OPD
HRP TMB-S
HRP-TMB
Alk phosphatase
B-galact ONPG
Urease
MTT
XTT
WST-1
Methylene Blue
Open assay 01
Open assay 02
Open assay 03
Open assay 04
Open assay 05
Open assay 06
Open assay 07
Open assay 08
Open assay 09
Open assay 10
Open assay 11
Open assay 12
Open assay 13
Open assay 14
Open assay 15
Open assay 16
Open assay 17
Open assay 18
Open assay 19
Open assay 20
Open assay 21
Open assay 22
Open assay 23
Open assay 24
Open assay 25
Figure D-8: Assay List
PowerWaveX Operator’s Manual
D-9
D-10
Sample Reports
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