Western Processor - Bio-Rad

Western Processor - Bio-Rad
Western
Processor
Instruction
Manual
Catalog Number
170-3970
For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call 1-800-4BIORAD (1-800-424-6723)
Safety
Caution/Warning
!
!
Disconnect power to the Western Processor before servicing. No user-serviceable parts
are inside the instrument. Refer servicing to Bio-Rad service personnel.
!
This instrument is intended for laboratory use only.
This product conforms to the “Class A” standards for electromagnetic emissions
intended for laboratory equipment applications. it is possible that emmissions from this
product may interfer with some sensitive applicances when placed nearby or in the same
circuit as those applicances. The user should be aware of this potential and take appropriate
measures to avoid interference.
The Western Processor is certified to meet the EN61010-1 safety standard for safety of
laboratory equipment. Certified products are safe to use when operated in accordance with the
instruction manual. This safety certification does not extend to other equipment or accessories not EN61010-1 certified, even when connected to the Western Processor.
This instrument should not be modified or altered in any way. Alteration of this instrument
will void the manufacturer’s warranty, void the EN61010-1 certification, and create a potential safety hazard for the user.
Bio-Rad is not responsible for any injury or damage caused by the use of this instrument
for purposes other than those for which it is intended or by modifications of the instrument not
performed by Bio-Rad or an authorized agent.
EN61010-1 is an internationally accepted electrical safety standard for laboratory instruments.
Warranty
The Western Processor is warranted for one (1) year against defects in materials and
workmanship. If any defects should occur during this warranty period, Bio-Rad Laboratories
will replace the defective parts without charge. However, the following defects are specifically
excluded:
1. Defects caused by improper operation.
2. Repair or modification done by anyone other than Bio-Rad Laboratories or their authorized agent.
3. Use with other spare parts not specified by Bio-Rad Laboratories.
4. Damage caused by deliberate or accidental misuse.
5. Damage due to use of improper solvent or sample including acetic acid and methanol.
For inquiry or request for repair service, contact your local Bio-Rad office.
Table of Contents
Section 1
Introduction ..................................................................................................1
1.1
1.2
1.3
1.4
1.5
Overview ....................................................................................................................1
Features.......................................................................................................................1
Unpacking...................................................................................................................1
System Set-up.............................................................................................................2
Description of System ................................................................................................3
Section 2
System Operation.........................................................................................4
2.1
2.2
Flowchart of Main Operating Modes.........................................................................4
Flowchart of Pause Subroutines.................................................................................5
Section 3
Mode I—Running a Programmed Method ..............................................7
3.1
3.2
3.3
Flowchart for Running a Programmed Method.........................................................8
Sequence of Steps to Run a Programmed Method ....................................................9
Sequence of Steps for Pause During Assay.............................................................10
Section 4
Mode II—Programming or Editing a Method .......................................11
4.1
4.2
4.3
Flowchart for Programming or Editing a Method...................................................11
Sequence of Steps to Program or Edit a Method.....................................................12
Sequence of Steps for Pause in Edit Mode..............................................................15
Section 5
Mode III—Calibrate Pump Mode ...........................................................16
5.1
5.2
Flowchart for Calibrate Pump Mode .......................................................................16
Sequence of Steps to Calibrate Pump ......................................................................17
Section 6
Mode IV—Purge Tubing Mode................................................................18
6.1
6.2
Flowchart for Purge Tubing Mode ..........................................................................18
Sequence of Steps to Purge Tubing .........................................................................19
Section 7
System Maintenance..................................................................................21
7.1
7.2
7.3
Pump Pressure Plate.................................................................................................21
Tube Cleaning ..........................................................................................................21
Tube Replacement....................................................................................................21
Appendix A Troubleshooting..........................................................................................22
Appendix B Pre-programmed Methods........................................................................23
Appendix C Product Information and Specifications .................................................29
Appendix D Worksheet for Programming a Method ..................................................31
Section 1
Introduction
1.1 Overview
The Western Processor automates the western blot development process. After the antigen
is applied to the membrane surface, the Western Processor will process the blot through the various washing, blocking, and incubation steps. Once the system is set-up with the required
reagents and the appropriate assay is programmed, the Western Processor will proceed with the
assay and alert you at certain critical steps and when the assay is completed.
The Western Processor is fully programmable from the front panel and can store up to
ten (10) user defined assays. Each assay holds up to fifteen (15) steps where each step can contain up to six (6) cycles. In addition to these ten (10) user defined assays, the Western Processor
contains five (5) pre-programmed assays designed specifically for use with Bio-Rad’s
immunoassay kits.
The Western Processor holds one, two, or four Mini-blot Trays for processing mini blots
and one Standard Tray for processing up to 20 x 20 cm blots.
1.2 Features
The Western Processor provides the following features:
•
Automatically controls incubation times, volumes, and washes.
•
Six (6) programmable reagent delivery and manual reagent delivery cycles.
•
Five (5) pre-programmed assays.
•
Store up to ten (10) user-defined assays.
•
Internal diaphragm pump aspirates reagents to waste bottle.
•
The optional reagent rack holds six 50 ml tubes and two 1 liter bottles.
•
By using the purge cycle, the tubing is easy to clean.
•
Barbed fittings allow for easy removal and replacement of tubing.
1.3 Unpacking
When you receive the Western Processor, carefully inspect the shipping containers for any
damage which may have occurred in shipping. Severe damage to a container may indicate damage to its contents. If you suspect damage to the contents may have occurred, immediately file a
claim with the carrier in accordance with their instruction before contacting Bio-Rad Laboratories.
Open the shipping carton and lift the contents out of its packing. Check the contents of the
box against the supplied packing list. Inspect the instrument for external damage. If any part
is missing or damaged, contact Bio-Rad laboratories immediately.
Packing List
Western Processor
Standard Tray with lid, 1
Mini-blot Trays, 4
Waste bottle with lid and fittings
Dispensing and aspiration tubing assemblies including 3 tubing kits for 3 tray modes
External power supply with power cord
Tubing labels
Instructions
1
1.4 System Set-Up
1. Level the Western Processor on the work surface where you intend to operate. Connect
the power cord to the external power supply and then plug the power supply into the back
of the unit.
2. Turn the Western Processor on by pressing the ON/OFF power switch on the front panel
of the unit.
3. The LCD displays the start-up screen with the software version, then the platform levels
to the horizontal home position.
4. Unwrap the Standard Tray with lid, four Mini-blot Trays, and tubing kits.
5. Remove the dispensing and aspiration tube assemblies and reverse lid.
6. Place Standard Tray with lid on the platform.
7. Choose a tubing kit appropriate for the experiment (one tray, two tray or four tray), each
of which has a dispensing tube assembly (smaller diameter tubing) and an aspiration
assembly (straws on the end).
8. Connect the open fitting of the aspiration assembly to the tubing connected to the waste
bottle cap. (When changing tubing kits for experiments with different numbers of trays,
disconnect and reconnect from the same junction, the end of the tube coming from the
waste bottle). Insert the straw end(s) into the 3 mm hole(s) above the tray positions you
will be using as in the photographs in Figure 1.
9. Connect the tubing from the waste outlet on the back panel of the Western Processor to
the second fitting in the waste bottle cap.
10. Connect the open fitting from the dispensing tube assembly to the 3mm tubing connected
at the other end to the Y fitting after the pump. Put each dispensing end in the 0.8 mm hole
in the lid next to each aspiration tube. Again refer to figure 1 for placement of the tubing
kits for each tray configuration.
Fig. 1a. 4 tray configuration.
Fig. 1b. 2 tray configuration.
2
Fig. 1c. 1 tray configuration.
1.5 Description of System
Front Panel of Western Processor
ENTER:
Use this key to select the message displayed on the LCD.
YES/», NO/«: These keys are used to scroll through the different options on the LCD and
to select or decline messages displayed on the LCD.
SILENCE
ALARM:
This key is used to silence the alarm.
• Reagent preparation/addition alarm. When the program includes a step
for manual addition of reagent, or the alarm is programmed in a step with
automatic reagent addition, the alarm will sound at two different time
points. The first alarm will sound five minutes prior to that step to alert the
operator to prepare the required reagent. The second alarm sounds
immediately prior to that step and prompts the operator to either prime
the necessary valve, or add the reagent manually.
• Variable incubation alarm. Valve 4/SUBST allows for programming a
variable incubation time. An alarm will sound at the end of the programmed
time to alert the operator to monitor and stop the development.
• End of assay alarm. An alarm will sound after the final step of a assay.
• Manual aspiration alarm. When a "no aspiration" step is programmed, an alarm
will sound to alert user to remove the reagent. This is generally used for a step
where the blot was incubated with a valuable reagent that may be re-used. The
platform continues rocking until the user reponds to the alarm. After the reagent
is removed, the user is prompted to push a key to continue the assay.
PAUSE:
During a run this key is used to temporarily stop the unit, this allows you to
make changes during the run, or return to the run (page 5). During
programming, this key can be used to access certain subroutines (page 6) or
to escape to the beginning of a programming step, then to the beginning of
a program.
ON/OFF:
The Western Processor is turned on by pressing the ON/OFF switch on the
front panel of the instrument.
The ON/OFF switch can also be used to reset the instrument. When resetting
the instrument in the edit mode, all steps are saved to memory.
3
Rear panel of Western Processor
waste line to waste bottle
Peristaltic Pump for
Six valves for reagent
Section 2
System Operation
The Western Processor is designed to provide simple, intuitive screen layouts for programming. The system consists of four main modes that allow the operator to, I) run a programmed assay, II) edit or program an assay, III) calibrate the pump, IV) and purge tubing.
2.1 Flowchart of Main Operating Modes
Mode I: Running a Method
Start a programmed assay.
READY FOR A NEW TEST
Mode II: Program/Edit a Method
Program a new method or edit a previously programmed method.
Mode III: Calibrate Pump Mode
Enter pump calibration mode. In this
mode the instrument will adjust the
“on” time of the pump to calibrate the
dispensed volume The instrument
comes factory-calibrated. Check the
calibration periodically and whenever
the pump tubing is replaced.
Mode IV: Purge Tubing Mode
Enter the manual or autoflush lines
mode.
ENTER EDIT MODE
CALIBRATE PUMP
PURGE TUBING
NO
4
2.2 Flowcharts of Pause Subroutines
In addition to the four main operating modes, two subroutines are accessible with the
PAUSE key.
PAUSE While Running a Method
This will let you make certain changes while running a method, such as priming valves,
increasing incubation times, skipping a step, and adding additional cycles to a step. By putting
the instrument in pause you have access to make these changes.
Note: Changes made while running a method are not permanent. When the method is
completed, it reverts to the originally programmed parameters.
Ready for a New Test
Start Assay?
Press PAUSE key during any step
01:
02:
03:
04:
05:
Return to assay
Prime Valves
Skip to Step
Extend Incubation
Additional Cycles
Option 01:
Option 02:
Option 03:
Option 04:
Option 05:
5
Restarts assay at the step it was paused.
Enters prime valve routine.
Allows you to skip a step during a method.
Increase the incubation of the paused step.
Increase the number of cycles of the paused
step.
PAUSE at the “Enter Edit Mode” Prompt
This will allow access to: 1) changing valve names, 2) changing the number of waste
bottles (1 or 2), and 3) changing the rocking speed of the platform. In this mode you can also
use the Western Processor as a simple rocking platform by turning the rocking motor on at a
specified speed.
Ready For a New Test
Enter Edit Mode
Waste Bottle(s)
Set Rocking Speed
Change Valve Names
Select 01 or 02 waste
bottles.
Set Rocking Speed
for Slow, Med, and
Fast Speeds. Use system as rocking table.
Allows operator to
change names for
valves.
Note: The Set Rocking Speed will cycle through FAST, MED, and SLOW speed modes.
The preset values for each of these rocking speeds are 30 RPM for FAST speed, 15 RPM
for MED speed, and 5 RPM for SLOW speed.
Caution: All programmed methods will reflect the new valve names.
6
Section 3
Mode I–Running a Programmed Method
When the Western Processor is turned on, the prompt "Ready for a New Test" appears
after the initial start up screen. By pressing the YES/» key, you enter a sequence of steps to
start a programmed method.
The Western Processor contains five (5) pre-programmed assays to process mini-blots
with Bio-Rad’s immunoassay kits. These pre-programmed assays are designed to produce
the optimum signal to noise ratio. The parameters of these pre-programmed assays cannot be
changed. However, a similar assay can be programmed that is specifically tailored to your
needs. For example, if you have a limited amount of primary antibody, you can reduce the primary antibody volume to as low as 5 ml. We recommend keeping the washing step volumes,
rocking speeds, and times as listed in these assays. See Appendix B for details of the five
assays.
The five pre-programmed assays and corresponding Bio-Rad immunoassay kit are:
#1–IBLOT:
Bio-Rad Immun-Blot Kit
#2–AMPAP:
Bio-Rad Amplified Alkaline Phosphatase Immun-Blot Assay Kit
#3–OP4CN:
Bio-Rad Opti-4CN Detection Kit
#4–AOPTI:
Bio-Rad Amplified Opti-4CN Detection Kit
#5–ISTAR:
Bio-Rad Immun-Star Chemiluminescent Protein Detection Systems
7
3.1 Flowchart for Running a Programmed Method
READY FOR A NEW TEST?
YES
NO
Assay Name
01: IBLOT
Enter Edit Mode
Pump Pad in Place?
Press YES
YES
Prime Valves?
NO
Tray Count
01 (02) 04
INSTALL TRAY(s)
Press Yes
Start Assay
YES
NO
START ASSAY
Positioning platform
Prime Line 01: WASH1
Prime Line 02: WASH2
Etc.
Press yes or no at
Aspirating
Prime Again?
YES
NO
Positioning Platform
Note: When a method is completed, be sure to immediately wash all six (6) lines and
valves in the Purge Tubing Mode. If buffer salts and other reagents are allowed to remain
in the system, the lifespan of the valves will be greatly reduced.
8
3.2 Sequence of Steps to Run a Programmed Method
Step #
01
LCD Display
Ready for a New Test
YES or NO
Description
YES: Enters step sequence to start a method
NO:
Enters Edit Mode
02
Assay Name
01: ------
Use YES/» and NO/« keys to scroll through methods
01 to 15. Methods 1 through 5 are pre-programmed
assays. Press ENTER to select the program displayed
on the LCD
03
Pump Pad in Place
PRESS YES
Make sure the pump pad on the back panel of the
instrument is in place.
Press YES to continue.
Pressure Plate
04
Prime Valves
YES or NO
05
Install Tray(s)
Press YES
06
Prime Line 01: WASH1
Prime Line 02: WASH2
Etc.
YES: Enters Prime Valves routine. This allows
the operator to prime selected valves prior to starting a
method.
NO:
Ready to start assay, go to step # 08 below
Place tray(s) on the platform prior to priming the
valves. One or two Mini-blot Trays, or one Standard
Tray. Make sure the delivery- and aspirating- tubes are
properly positioned in the lid. Press YES to continue.
The platform will automatically position itself.
YES: Primes that line
NO:
Advance to the next line. You can also prime
the lines during the assay itself using the Pause
Mode.
YES or NO
07
Prime Again?
YES or NO
08
Tray Count
01 (02) 04
09
Start Assay
YES or NO
Each valve will be sequentially displayed on the
LCD. When all 6 lines are processed, the system will
automatically aspirate the dispensed reagents.
YES: Repeat the priming sequence of step 06.
NO:
Ready to start assay. The platform will automatically position itself.
Use YES/» and NO/« keys to select 01, 02, or 04
trays.
ENTER: Selects number displayed on LCD.
(Selecting the number of trays will inform the Western
Processor to deliver and aspirate the required volumes.)
YES: Starts the selected Method
Before pressing yes, make sure all the reagents are in
place, and the necessary lines have been primed. Lines
for reagents that are prepared just before use, can be
primed in the pause mode during the assay.
NO:
Returns to “READY FOR A NEW TEST”
mode.
9
Note: When a method is completed, be sure to immediately wash all six (6) lines and
valves in the Purge Tubing Mode. If buffer salts and other reagents are allowed to remain
in the system, the lifespan of the valves will be greatly reduced.
4.3 Sequence of Steps for Pause During Assay
While running a method the system can be paused to prime valves, to skip a step in the
method, to extend the incubation time, or to add additional cycles to the paused step. To
access these features, you can enter this subroutine any time during a method by pressing the
PAUSE key.
Note: Changes made while running a method are not permanent. When the method is
completed, the method reverts to the originally programmed parameters.
Step #
01
02
03
04
05
05a
LCD Display
Step #: Reagent
Cycle # Time
Pause Mode 01:
Return to Assay
Pause Mode
01: Return to assay
Pause Mode
02: Prime Valves
Pause mode
03: Skip to Step?
Skip to Step?
#: Reagent
06
Pause Mode
04: Extend Incubation
06a
Extend Incubation
time by 01 min.
07
Pause Mode
05: Additional Cycles
Step#: Reagent
Total Cycles 04
07a
Description
Press PAUSE to access special subroutine.
Use the YES/» and NO/« keys to scroll through
the following five options:
01: Return to Assay
02: Prime Valves
03: Skip to Step
04: Extend Incubation
05: Additional Cycles.
Press ENTER to select the option
ENTER: Return to assay
ENTER: Enters the prime valve routine. Before
starting the prime valve sequence, remove both
dispense lines from the lid and place them in a
container. Follow on screen prompts.
ENTER: Enters skip step routine.
Use YES/» and NO/« keys to choose the step
number. Press ENTER to select the step. Method
proceeds with the selected step.
ENTER: Enters incubation time routine. You can
extend the incubation time of the paused step by
sixty (60) minutes.
Use YES/» and NO/« keys to enter the time. The
incubation time of the paused step will be extended by the time entered here. Press ENTER to select
the entered time. Method proceeds where paused.
ENTER: Enter additional cycles to routine
Use YES/» and NO/« keys to enter the number of
cycles (01 to 06). The maximum number of cycles
in a single step is six. Press ENTER to select the
total number of added cycles. Method proceeds
where paused.
10
Section 4
Mode II—Programming or Editing a Method
In this mode you can program a new method or edit an existing method. You can program
up to 10 customized methods, each method consisting of up to 15 steps. Each step allows
programming for reagent, incubation time, volume, rocking speed, and number of cycles. In
addition, with subst/valve#4 reagent you can program a variable incubation time that alerts
the operator to monitor the development step after a specified time.
4.1 Flowchart for Programming or Editing a Method
Ready for a New Test
YES
NO
ENTER EDIT MODE
YES
NO
Calibrate Pump
Mode
Edit Program
01: Empty
Change Program Name
YES
NO
Assay Name
#: --Assay Name - ...... OK?
YES
NO
Edit Step
01: EMPTY
Step # -- Reagent?
EMPTY (or Reagent & valve #)
When editing
existing method
Reagent INCUBATION
00 Hr
01 min
Reagent VOLUME
025 ml.
Reagent Rocking speed
MED
When programming
steps for a new
method.
Reagent Alarm?
YES
STEP # : Reagent
# of cycles 01
Aspirate after this step
YES
Edit another step?
YES
11
NO
Note:
The pause key can be used
to back up one level if a
mistake is keyed in. From
within a step, pause backs
up to the beginning of that
step (EDIT STEP _). From
the beginning of a step,
pause backs up to ENTER
EDIT MODE.
4.2 Sequence of Steps to Program or Edit a Method
Note: Pause key can be used during programming to back up.
Step #
LCD Display
Description
01
Enter Edit Mode
YES: Enters programming or edit mode,
YES or NO
go to step 02
NO:
02
Edit Program
01: Empty
Enters Pump Calibration Mode
Use the YES/» and NO/« keys to scroll through the
list of programmed methods. Press ENTER to select
the program displayed on the LCD screen. The program name "EMPTY" indicates a new method can be
programmed. Selecting a previously programmed
method allows you to edit any or all steps in that
method.
Note: Programs 1 through 5 are pre-programmed
assays. No changes can be made to these assays. See
Appendix B for details of these five assays.
03
Change Program Name
YES or NO
YES: Name a new program or change the name of
a previously programmed method.
NO:
Retain the displayed name, go to step # 6.
04
Assay Name
#:—
Use the YES/» and NO/« keys to scroll through the
alphabet and numbers. 012345689ABC….XYZ. The
LCD screen displays a cursor under the space for the
first character. Scroll to choose the selected character
then press ENTER to select the character. The cursor
moves to the next space for the following character.
Each name can consist of up to 5 characters.
05
Assay Name ——-OK?
YES or NO
YES: Saves displayed method name.
NO:
Rejects displayed method name, returns to
step 04 to re-enter the name.
06
Edit Step
01: EMPTY
Press ENTER when programming a new method to
enter step 1 of the method.
When editing a method,
the display will show
the actual reagent name
in place of EMPTY.
When editing a method, use the YES/»
and NO/« keys to scroll through the
different steps, then press ENTER to select the step
you wish to edit.
STEP # REAGENT?
EMPTY (or Reagent
and valve#)
Use the YES/» and NO/« keys to scroll through the
different choices and press ENTER to accept the
reagent/valve# choice.
07
WASH1
WASH2
CONJ1
SUBST
Valve 1
Valve 2
Valve 3
Valve 4 (allows for variable incubation time)
CONJ2
Valve 5
REAGT
Valve 6
MANUL
(Allows for manual addition of
reagent)
SKIP Can be used to hide a step while editing a
method
END Returns to Enter Edit Mode prompt.
12
Step #
08a
LCD Display
Description
Reagent INCUBATION Use the YES/» and NO/« keys to enter the number of
00 HR 01 MIN
hours for the reagent incubation. Press ENTER to
select the entered number of hours. Then use the
(When editing, the
YES/» and NO/« keys to enter the number of minutes
display will show the
for reagent incubation. Press ENTER to select the
programmed reagent
entered number of minutes.
name)
08b
Variable Incubation
YES
Use the YES/» and NO/« keys to choose YES or NO,
then press ENTER to select the choice.
Variable incubation is available only when
SUBST/Valve 4 is selected. When variable incubation
is selected, an alarm will sound at the end of the programmed incubation time. Incubation will continue
until you press NO at the prompt. This allows you to
monitor band development and stop the reaction when
the bands are sufficiently developed.
9
Reagent VOLUME
025 ml
Use the YES/» and NO/« keys to enter the
volume of reagents to be dispensed, from 000 ml to
500 ml in 1 ml increments. Press ENTER to select
the number entered.
The volume entered is per tray. When starting a
method, you will be prompted to enter the number of
trays used. The instrument will automatically dispense
the programmed volume in each tray.
When entering a MANUL step where reagent is manually added, you are prompted to enter the volume of
the reagent you will add. (See Step 14)
10
Reagent Rocking Speed
MED
Use the YES/» and NO/« keys to scroll through the
Rocking Speed options. The Rocking Speed options
are, ZERO, SLOW (5 rpm), MED (15 rpm), and FAST
(30 rpm). These are the pre-set values. We recommend
Medium–Slow Speed for incubation steps and Fast
speed for wash steps. To change the pre-set values enter
PAUSE subroutine in Enter Edit Mode.
13
Step #
11
LCD Display
Reagent Alarm?
YES
Description
Use the YES/» and NO/« keys to choose Alarm, then
press ENTER to select. If the step you define requires
freshly prepared reagent, you can set the alarm to
prompt you to prepare the reagent. When the alarm is
selected, the alarm will first sound five minutes before
the step that requires the freshly prepared reagent. A
second alarm will sound when it is time to add the
reagent. The instrument display will prompt you to
prime the valve.
12
Step # Reagent
# of cycles 01
Use the YES/» and NO/« keys to choose 01 to 06
cycles in that step, then press ENTER to select.
13A
Aspirate After This
Step?
YES
Use the YES/» and NO/« keys to select or deselect
aspirate, then press ENTER to select. When a single
cycle is selected in a step you can select to aspirate that
step. If NO is selected, the instrument will not aspirate
the reagent and the reagent for the next step be added to
the tray(s).
When multiple cycles are selected the instrument will
automatically aspirate after each cycle except when it is
the final step in a method. In that case the last cycle of
that step will not be aspirated to prevent drying out of
the blot(s).
13B
If two (2) waste bottles were selected in the Edit Mode
Pause subroutine, you will now be asked to assign
waste bottle 01 or 02 with each step. If waste bottle 02
is selected an alarm will sound upon completion of that
step to manually transfer the waste lines to the second
waste bottle, or manually remove reagent. For example,
to recover primary antibody, selecting waste bottle 2
allows you to recover the primary antibody.
14
MANUL Asp Volume?
25 ml
This display appears only when you have selected
MANUL for reagent. Entering the volume of the
reagent you will manually add per tray alerts the instrument to the volume to be aspirated. Use the YES/» and
NO/« keys to choose volume (000 to 500ml) then press
ENTER to select. Reagent can be recovered by using
the 2 waste bottle option.
15
Edit Another Step?
YES or NO
YES: Return to Step 7 in this programming
sequence to program another step.
NO:
14
Return to “ENTER EDIT MODE” screen.
4.3 Sequence of Steps for Pause in Edit Mode
A separate subroutine can be accessed through the ENTER EDIT MODE screen. This subroutine allows the operator to set the number of waste bottles, to set the rocking speed, and to
change the valve names.
Step #
01
LCD Display
Enter Edit Mode
YES
NO
Description
Press PAUSE to enter subroutine
02
Waste Bottle(s)
01
Use YES/» and NO/« keys to enter 01 or 02 waste
bottles. When one waste bottle is selected all reagents
will be aspirated into this single container. Select two
waste bottles if you wish to retain certain reagents.
When two waste bottles are selected, you will be asked
at each step during programming whether the reagent
will be aspirated in waste bottle 1 or 2.
03
Set Rocking Speed
YES
NO
YES:
NO:
Enters Set "Fast Rocking Speed:"
Enters Change Valve Names–go to step 8
04
Set Fast Rocking?
YES
NO
YES:
NO:
Enter screen to set Fast Rocking Speed.
Enters Set Med Rocking Speed.
05
Fast Rock
RPM=030
Use YES/» and NO/« keys to scroll to the desired
rocking speed (05 to 035 RPM). Press ENTER to select
the desired speed.
06
Run Rock Motor
YES
NO
YES:
NO:
07
Rock Motor On
Press NO to stop
This mode lets you use the Western Processor as a
standard rocking platform. Press NO to stop the motor
and return to "Set Fast Rocking" screen
Starts rocking motor at set RPM.
Return to Set Fast Rocking screen-go to step 4
Note: The Set Rocking Speed will cycle through FAST, MED, and SLOW speed
modes. The preset values for each of these rocking speeds are 030 RPM for FAST
speed, 015 RPM for MED speed, and 005 RPM for SLOW speed.
08
Change Valve Names
YES
NO
YES:
NO:
Enters Changing Valve Name routine
Returns to "Enter Edit Mode" screen
09
Valve 01: WASH1
Change? YES NO
YES:
NO:
Allows user to change name of Valve 01
Advances to Valve 02
10
Valve Name
01: ——-
Use YES/» and NO/« keys to scroll through alphabet
and numbers. The name can consist of up to
5 characters. After entering each character press
ENTER to select that character and advance to the next
character. Returns to "Enter Edit Mode" screen when
completed.
Note: The routine will scroll through all six valves. You can rename any of the valves.
The default valve names are: #1 WASH1, #2 WASH2, #3 CONJ1, #4 SUBST, #5
CONJ2, #6 REAGT.
Caution: All programmed methods will reflect the new valve names.
15
Section 5
Mode III—Calibrate Pump Mode
Pump calibration adjusts the time necessary for the pump to deliver a certain volume.
The Pump Calibration mode is accessed by pressing the NO/« keypad at the "ENTER EDIT
MODE?" prompt. The instrument comes factory calibrated. However, calibration should be
checked for accuracy periodically and when the pump tubing is replaced. If the dispensed
volume is incorrect, check that the tubing and nozzles are clean and in good condition. To
calibrate the dispensed volume, use the Pump Calibration Routine below.
5.1 Flowchart for CALIBRATE PUMP MODE
READY FOR A NEW TEST
YES
NO
Enter Edit Mode
YES
NO
CALIBRATE PUMP
YES
NO
Exercise New Tubing?
YES
NO
Pump pad in place
Press YES
Breaking in
tubing (20 min.)
Calibrate Now?
YES
NO
Dispense 40 ml now?
YES
NO
Volume Dispensed
040 ml
Calibration Complete
16
Purge tubing
YES
NO
5.2 Sequence of Steps to Calibrate Pump
Before calibrating the pump, make sure the pressure pad on the rear panel of the instrument has been locked into place.
Step #
01
LCD Display
Calibrate Pump?
YES NO
Description
YES: Enter Pump Calibration Routine
NO:
Enter Purge Tubing Routine
02
Exercise New Tubing?
YES NO
YES: Exercises new tubing (~20 minutes). LCD
Display will prompt you to make sure the pressure pad
is in place. The tubing supplied with the instrument
does not require the exercise routine. This routine is
used for new tubing. DO NOT use any fluid in the system during the exercise routine.
NO
Enter Pump Calibration (skip exercise tubing
routine).
03
04:
Calibrate Now?
YES NO
Dispense 40 ml Now?
YES NO
Before calibrating the pump make sure the pressure pad
has been locked into place for at least one hour in order
to approximate real-life operating conditions. It is also
best to prime the pump before starting calibration.
YES:
Continue Calibrate Pump Routine.
NO:
Return to Calibrate Pump Mode.
Remove dispense tube from dispense tubing at fitting
juction and place in a 50 ml graduated cylinder.
Make sure the valve 1 feed line is placed in water before
pressing YES.
YES:
NO:
05:
Volume Dispensed
40 ml
The pump will start and deliver 40 ml.
Return to Calibrate Pump Routine
ENTER : Volume delivered is 40 ml, return to
Calibrate Pump Mode.
If volume delivered is other than 40 ml, use the YES/»
and NO/« keys to enter the actual volume delivered,
then press ENTER. Return to step 03, Calibrate Now,
to verify calibration.
17
Section 6
Mode IV—Purge Tubing Mode
The tubing and valves should be purged at the completion of each assay to insure the
lines are free of contamination before the next assay. To remove any latent bacteria we recommend using a 2% bleach solution or a commercially available cleaning preparation that will
not interfere with your assay(s) or damage the tubing.
6.1 Flowchart for Purge Tubing Mode
Ready For a New Test
Enter Edit Mode
Calibrate Pump
PURGE TUBING
AUTOFLUSH LINES
Install Tray(s)
Install Tray(s)
Put Tubes in Cleaner
Flush Tubing
Positioning Platform
Dispensing Cleaner
Positioning Platform
Dispensing Reagent
Flush Tubing Again?
Put tubes in DI
Dispensing DI Water
Soaking tubes 05 min
Dispensing DI Water
Remove Tubes from DI
PRESS ENTER
Drying Tubes
Positioning Platform
Release Pressure pad
PRESS ENTER
18
Return to Ready
for a New Test
screen
6.2 Sequence of Steps to Purge Tubing
For optimum performance we recommend utilizing the purge tubing routine after each
assay. During the Purge Tubing Mode, the tubing from each valve is cleaned. The Purge
Tubing Mode is accessed via the Pump Calibration routine or at the completion of an assay.
Below are the step by step detailed instructions for purging the tubing.
Step #
01
LCD Display
Purge Tubing?
YES NO
Description
YES: Enter Purge Tubing Routine
NO:
Return to Ready for New Test
02
Autoflush Tubes?
YES NO
YES: Enters sequence of three flushing
cycles, 1) wash and soak, 2) rinse and soak, and 3)
dry. GO TO STEP 03 then 06.
NO:
Enters sequence of steps for single wash
cycles. FOLLOW STEPS 03 and 04.
03
Install Tray(s)
Press YES
Make sure a tray is on the platform, then
press YES. Make sure the two dispense lines and the
two aspirate lines are positioned over the tray(s).
Make sure the six tubes are placed in the appropriate
wash solution.
04
Manual
Flush
Mode
Flush Tubing?
YES NO
YES: Enter manual flush tubes routine. Platform
positions itself automatically.
NO:
Return to Purge Tubing screen.
05
Dispensing
Valve 1, 2, 3,....6
The instrument will sequentially wash each of the six
tubes. After completion, the dispensed solution will
be aspirated and the routine enters "Flush Tubing
Again" to repeat washing/rinsing cycle or to return to
Purge Tubing Screen.
06
Put Tubes in Cleaner
Autoflush Press ENTER
Mode
Place the six tubes in the appropriate cleaning solution, then press ENTER.
07
Dispensing Cleaner
Valve 1, 2, 3,....6
The instrument will sequentially pump cleaning
solution through each of the six tubes. After completion the dispensed solution will be aspirated.
08
Soaking Tubes 05 min
The tubes will soak for 05 minutes in the cleaning
solution. After 5 minutes the alarm will sound.
09
Put tubes in DI
Press ENTER
Place the six tubes in a container with deionized
water and press ENTER.
10
Dispensing DI water
Valve 1, 2, 3,....6
The instrument will sequentially pump deionized water
through each of the six tubes. After completion the
dispensed solution will be aspirated.
11
Soaking Tubes 05 min
12
Dispensing DI water
Valve 1, 2, 3,....6
The tubes will soak for 05 minutes in di water. A second wash will automatically start, step 12.
For the second wash, the instrument will sequentially
pump deionized water through each of the six tubes.
After completion the dispensed solution will be aspirated. The alarm will sound to alert you to step 13.
19
Step #
13
LCD Display
Remove tubes from DI.
Press ENTER
Description
Remove the tubes from the di water.
ENTER: Starts pump to remove remaining deionized
water and air flush the tubes.
14
Drying tubes 05 min.
Valve 1, 2, 3,....6
15
Positioning Platform
Platform will automatically position itself.
16
Release Pressure Pad
Press ENTER
Unlock the pump pressure pad from the pump on the
rear panel of the instrument. ENTER: Returns to Ready
for a New Test screen.
20
Section 7
System Maintenance
7.1 Pump Pressure Plate
The spring loaded pressure plate on the peristaltic pump should be unlocked when the
instrument is not in use to minimize wear on the tubing.
Pressure Plate
The tension setting on the pump release is preset for optimum performance. Changing the
tension of the pump release will adversely affect the calibration and the lifespan of the tubing.
7.2 Valve and Tube Cleaning
The valves and tubing should be kept clean to ensure good pumping action. Upon completion of an assay the operator is given the opportunity to purge the tubing. You can choose
to purge the tubing in the manual or automatic mode. See Section 6 for detailed steps on the
Purge Tubing Mode. To clean the tubing, we recommend you use a 2% bleach solution or a
commercially available preparation to remove any latent bacteria. In the automatic purge
tubing mode, the lines are soaked for five minutes with the cleaning solution. Subsequent
washes remove the cleaning solution and dissolve any accumulation of salt.
7.3 Tube Replacement
Tube replacement kits are available, see Appendix C for product ordering information.
The tubing in these kits is cut to the proper length. Proper tube length and position are
critical for proper performance of the peristaltic pump. The pump tubing has an approximate
life of 1,000 hours of pumping. Pumping occurs for a fraction of assay time and is dependent
on volumes delivered. If you notice liquid on the bench - change the pump tubing. It is a good
idea to have a set of replacement pump tubing available in your lab.
21
Appendix A
Troubleshooting
Problem
Reagents are not dispensed
Cause
1. Tubing is not connected.
2. Reagent bottle(s) are
empty.
LCD display does not light
up when instrument
is turned on
3. Pump Pressure Pad is in
unlocked position
Instrument not plugged into
the power pack and/or wall
outlet
Splashing during dispense
cycle
Tubes and/or nozzles are
plugged
"Rock Motor Steps Lost"
display error
Rock Motor fails to rock
platform correctly.
Instrument fails to aspirate
dispensed reagents.
1. Waste bottle not sealed.
2. Aspiration pump not
working properly.
3. No vacuum.
22
Solution
1. Connect tubing.
2. Make sure appropriate
reagent bottles are
filled.
3. Place Pump Pressure
Pad in locked position.
Make sure the instrument
is plugged into the power
supply and into the electrical outlet.
Clean or change tubing and
nozzle. If tubing is changed
proceed through the exercise tubing and pump calibration routine.
1. If the platform is still
rocking. Make sure
there is nothing impeding the movement of the
rocker mechanism.
2. If the platform is no
longer rocking. Call
Bio-Rad to return instrument for service.
1. Make sure the waste
bottle fittings are in
place and the lid is
secure.
2. If the aspiration pump
is not functioning call
Bio-Rad to return instrument for service.
3. Check for vacuum at the
hose extending from the
back of the instrument.
If vacuum is present at
the hose, change out
tubing form this point to
the bottle and from the
bottle to the aspiration
tip.
Problem
Unsatisfactory results.
E.g. high background.
Cause
1. Ran incorrect assay.
2. Washing steps missing
or inadequate.
3. Inactive reagents
23
Solution
1. Make sure the correct
assay is selected and
programmed.
2. Increase the number of
cycles, volume, or rocking speed of the wash
steps.
3. Check the manual for
the assay used to check
the reagents used.
Appendix B
The Western Processor contains five (5) pre-programmed assays to process mini-blots.
The parameters for each assay are listed below. These pre-programmed assays can be used as
templates for programming new assays to process large blots. Simply increase incubation volumes to 50 ml and wash volumes to 100–200 ml. Large blots may require a larger waste bottle,
which can be obtained from Nalgene Inc. (Heavy Duty Vacuum Bottle, catalog number 21264000, and Filling/Venting Closure, catalog number 2162-0831)
Assay # 1: IBLOT
The Immun-Blot assay kits are enzyme immunoassays kits optimized for the detection of
specific antigens. The kits provide all necessary components and chemicals. This pre-programmed assay is designed to produce the optimum signal to noise ratio. The parameters listed for each step cannot be changed in this pre-programmed assay. However, a similar assay
can be programmed that is specifically tailored to your needs. For example, if you have a
limited amount of primary antibody, you can reduce the primary antibody volume in step 4
to as low as 5 ml. We do recommend keeping the washing step volumes, rocking speeds, and
times as listed in this assay.
Note: When using the Protein G-HRP, the unbound primary antibody is removed with
TCBS pH 5.5, rather than TTBS pH 7.5. The optimal binding pH of Protein G is 5.5.
This assay can be used by manually exchanging the buffer at step 5, or by programming
a new assay.
Reagents Required
Valve #1 - WASH1:
TTBS, Tris buffered saline, 20 mM Tris, 500 mM NaCl, 0.05%
Tween-20, pH 7.5
Valve #2 - WASH2:
DI water
Valve #3 - CONJ1:
Primary antibody
Valve #4 - SUBST:
Alkaline phosphatase- or horseradish peroxidase color development reagents
Valve #5 - CONJ2:
Secondary antibody conjugate
Valve #6 - REAGT:
Blocking solution
Step #
1
2
3
4
5
6
7
8
9
10
Valve #/
Name
1: WASH1
6: REAGT
1: WASH1
3: CONJ1
1: WASH1
5: CONJ2
1: WASH1
4: SUBST
2: WASH2
End
Reagent
# Cycles/
Time
Volume/
Speed
Alarm/Aspirate/
Variable Incubation
TTBS
Blocking solution
TTBS
Primary antibody
TTBS
Secondary antibody
TTBS
developer
diH2O
1 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
4 x 5 min.
> 10 min
2 x 5 min
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/Yes
No/No/NA
Note: In step 8 an alarm will sound after 10 minutes incubation with substrate. At this
point, you can elect to stop or continue the development process.
24
Assay # 2: AMPAP
The Amplified Alkaline Phosphatase Immun-Blot assay kit is an enzyme immunoassay
system utilizing biotin-streptavidin for increased sensitivity in detection of specific antigens.
The kit provides all necessary components and chemicals. This pre-programmed assay is
designed to produce the optimum signal to noise ratio. The parameters listed for each step cannot be changed in this pre-programmed assay. However, a similar assay can be programmed
that is specifically tailored to your needs. For example, if you have a limited amount of primary antibody, you can reduce the primary antibody volume in step 4 to as low as 5 ml. We
do recommend keeping the washing step volumes, rocking speed, and times as listed in this
assay.
Reagents Required
Valve #1 - WASH1:
TTBS, Tris buffered saline, 20 mM Tris, 500 mM NaCl, 0.05%
Tween-20, pH 7.5
Valve #2 - WASH2:
DI water
Valve #3 - CONJ1:
Primary antibody
Valve #4 - SUBST:
Alkaline Phosphatase color development reagents
Valve #5 - CONJ2:
Secondary antibody - alkaline phosphatase conjugate.
Valve #6 - REAGT:
Blocking solution
MANUL:
Streptavidin-biotinylated alkaline phosphatase complex. When
preparing this reagent allow the complex to incubate for 1–3 hours
before using. If allowed to form over 3 hours, remake the solution.
See manual for details.
Step #
1
2
3
4
5
6
7
8
9
10
11
12
Valve #/
Name
1: WASH1
6: REAGT
1: WASH1
3: CONJ1
1: WASH1
5: CONJ2
1: WASH1
MANUL
1: WASH1
4: SUBST
2: WASH2
End
Reagent
# Cycles/
Time
Volume/
Speed
Alarm/Aspirate/
Variable Incubation
TTBS
Blocking solution
TTBS
Primary Ab
TTBS
Secondary Ab
TTBS
Streptavidin-AP
TTBS
developer
diH2O
2 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
4 x 5 min
≥ 10 min
2 x 5 min
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
Yes/Yes/NA
No/Yes/NA
No/Yes/Yes
No/No/NA
Note: In step 10 an alarm will sound after 10 minutes incubation with substrate. At this
point you can elect to stop or continue the development process.
25
Assay # 3: OP4CN
The Opti-4CN substrate kit is a more sensitive version of the colorimetric horseradish
peroxidase (HRP) Immun-Blot kit. The substrate, 4-chloro-1-naphthol (4CN) is replaced with
Opti-4 CN substrate producing a 4–8 fold increase in detection sensitivity. The parameters listed for each step cannot be changed in this pre-programmed assay. However, a similar assay
can be programmed that is specifically tailored to your needs. For example, if you have a
limited amount of primary antibody, you can reduce the primary antibody volume in step 4
to as low as 5 ml. We do recommend keeping the washing step volumes, rocking speed, and
times as listed in this assay.
Reagents Required (See manual for details of reagent preparation)
Valve # 1 - WASH1:
PBST, Phosphate buffered saline / 0.1% Tween-20
Valve # 2 - WASH2:
DI water
Valve # 3 - CONJ1:
Primary antibody
Valve # 4 - SUBST:
Opti-4 CN substrate
Valve # 5 - CONJ2:
Secondary antibody-HRP conjugate
Valve # 6 - REAGT:
Blocking solution
Step #
1
2
3
4
5
6
7
8
9
10
Valve #/
Name
1: WASH1
6: REAGT
1: WASH1
3: CONJ1
1: WASH1
5: CONJ2
1: WASH1
4: SUBST
2: WASH2
End
Reagent
# Cycles/
Time
Volume/
Speed
Alarm/Aspirate/
Variable Incubation
PBST
Blocking solution
PBST
Primary Ab
PBST
Secondary Ab
PBST
Opti-4CN Substrate
diH2O
2 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
3 x 5 min
1 x 60 min
4 x 5 min
≥ 10 min
2 x 5 min
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
20 ml/Med
40 ml/Fast
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
Yes/Yes/Yes
No/No/NA
Note: In step 8 an alarm will sound after 10 minutes incubation with substrate. At this
point, you can elect to stop or continue the development process.
26
Assay # 4: AOPTI
The Amplified Opti-4 CN enzyme immunoassay kit results in an additional 4–8 fold signal amplification compared to the Opti-4 CN enzyme immunoassay kit. In addition to replacing the 4-chloro-1-naphthol (4CN) substrate with Opti-4 CN substrate the amplified Opti-4 CN
kit uses streptavidin-HRP to further amplify the signal. The parameters listed for each step cannot be changed in this pre-programmed assay. However, a similar assay can be programmed
that is specifically tailored to your needs. For example, if you have a limited amount of primary antibody, you can reduce the primary antibody volume in step 4 to as low as 5 ml. We
do recommend keeping the washing step volumes, rocking speed, and times as listed in this
assay.
Reagents Required (See manual for details of reagent preparation)
Valve # 1 - WASH1:
PBST, Phosphate buffered saline / 0.1% Tween-20
Valve # 2 - WASH2:
20% DMSO/PBST
Valve # 3 - CONJ1:
Primary antibody
Valve # 4 - SUBST:
Opti-4 CN substrate
Valve # 5 - CONJ2:
Secondary antibody-HRP conjugate
Valve # 6 - REAGT:
Blocking solution
Additional reagents for manual addition at steps 8 and 11:
Step 8
- MANUL:
Step 11 - MANUL:
Step #
1
2
3
4
5
6
7
8
9
10
11
12
13
14
Bio-Rad Amplification Reagent (BAR), see manual for preparation
Streptavidin-HRP reagent
Valve #/
Name
Reagent
# Cycles/
Time
Volume/
Speed
1: WASH1
6: REAGT
1: WASH1
3: CONJ1
1: WASH1
5: CONJ2
1: WASH1
MANUL
2: WASH2
1: WASH1
MANUL
1: WASH1
4: SUBST
End
PBST
2 x 5 min
40 ml/Fast
Blocking solution
1 x 60 min
20 ml/Med
PBST
3 x 5 min
40 ml/Fast
Primary Ab
1 x 60 min
20 ml/Med
PBST
3 x 5 min
40 ml/Fast
Secondary Ab
1 x 60 min
20 ml/Med
PBST
4 x 5 min
40 ml/Fast
BAR
1 x 10 min
20 ml/Med
20% DMSO/PBST
4 x 5 min
40 ml/Fast
PBST
3 x 5 min
40 ml/Fast
Streptavidin-HRP
1 x 30 min
20 ml/Med
PBST
3 x 5 min
40 ml/Fast
Opti-4CN Substrate
≥ 10 min
20 ml/Med
Transfer manually to diH2O to stop rxn
Alarm/Aspirate/
Variable Incubation
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
No/Yes/NA
Yes/Yes/Yes
No/Yes/NA
No/Yes/NA
Yes/Yes/NA
No/Yes/NA
Yes/Yes/Yes
Note: In step 13 an alarm will sound after 10 minutes incubation with substrate. At this
point you can elect to stop or continue the development process.
27
Assay # 5: ISTAR
The Immun-Star chemiluminescent detection system is a non-isotopic method for immunodetection of specific antigens immobilized on nitrocellulose or PVDF membrane. This system uses the CDP-Star chemiluminescent substrate and enhancer which is activated by an
alkaline phosphatase enzyme conjugate. The assay, ISTAR, processes the blot to the development step. At that point the blot is removed from the Western Processor and development
is carried out as described in the manual. The parameters listed for each step cannot be changed
in this pre-programmed assay. However, a similar assay can be programmed that is specifically tailored to your needs. For example, if you have a limited amount of primary antibody,
you can reduce the primary antibody volume in step 4 to as low as 5 ml. We do recommend
keeping the washing step volumes, rocking speed, and times as listed in this assay.
Reagents Required (See manual for details of reagent preparation)
Valve # 1 - WASH1:
TTBS, Tris buffered saline/0.1% Tween-20
Valve # 2 - WASH2:
TBS, Tris buffered saline
Valve # 3 - CONJ1:
Primary antibody
Valve # 4 - CONJ2:
Secondary antibody-HRP conjugate
Valve # 5 - REAGT:
Blocking solution
Step #
1
2
3
4
5
6
7
8
Valve #/
Name
# Cycles/
Time
Reagent
Volume/
Speed
Alarm/Aspirate/
Variable Incubation
2: WASH2 TBS
2 x 5 min
40 ml/Fast
No/Yes/NA
5: REAGT Blocking solution
1 x 60 min
20 ml/Med No/Yes/NA
1: WASH1 TTBS
3 x 5 min
40 ml/Fast
No/Yes/NA
3: CONJ1
Primary Ab
1 x 60 min
20 ml/Med No/Yes/NA
1: WASH1 TTBS
3 x 5 min
40 ml/Fast
No/Yes/NA
4: CONJ2
Secondary Ab
1 x 60 min
20 ml/Med No/Yes/NA
1: WASH1 TTBS
4 x 5 min
40 ml/Fast
No/Yes/NA
End
Transfer manually to diH2O to stop rxn
Remove membrane from wash and proceed manually with development.
Development of the membrane is done manually on a piece of saran wrap. After 5 minutes
it is placed in a bag or in saran wrap to prevent membrane from drying out. Depending on the
type of membrane and developer used, the film is exposed anywhere from 1–10 minutes or
12–24 hours. See manual for details.
28
Appendix C
Product Information and Specifications
Catalog
Number
Description
170-3970
Western Processor, 100–240 VAC, Western Processor unit with
integral rocking platform, user interface, peristaltic and diaphragm
pump. Also includes standard tray with clear plastic lid, waste bottle,
four mini-blot trays , 3 aspiration and dispense tubing kits for one, two
or four tray configurations, manual
170-3973
Reagent Rack, holds two 1-liter bottles of wash buffer and six
reagent vials
170-3974
Mini-blot Trays, 10, 8.5 cm x 10 cm
170-3975
Mini-blot Trays, 50, 8.5 cm x 10 cm
170-3976
Standard Trays, 2, 21.5 cm x 21 cm
170-3972
Acrylic Lid, Standard Tray, 1
170-3978
Waste Bottle, includes lid and fittings
170-3980
Tubing kit; semi complete. Reagent through dispense connection,
includes all tubing and connectors for complete replacement from
reagent tubes, tubing connected to valves, through the dual channel
pump, to the connection with aspiration and dispense kits (170-3982,
170-3983, and 170-3984)
170-3981
Peristaltic Pump Tubing Kit, includes four dual tubing assemblies
with snap in pump fittings and through Y connectors on either end
170-3982
4 Tray Aspiration and Dispense Kit, includes four each of
aspiration and dispense tubing and fittings through the connector to
the tubing kit (170-3980). This kit connects to four holes in the tray lid
for each dispense and aspiration into four mini trays
170-3983
2 Tray Aspiration and Dispense Kit, includes four each of
aspiration and dispense tubing and fittings through the connector to
the tubing kit (170-3980). This kit connects to two holes in the tray lid
for each dispense and aspiration into two mini trays or large tray
170-3984
1 Tray Aspiration and Dispense Kit, includes four each of
aspiration and dispense tubing and fittings through the connector to
the tubing kit (170-3980). This kit connects to one hole in the tray lid for
dispense and aspiration into one mini tray
100-1547
Western Processor Service Manual
29
Specifications
Parameter
Number of trays
Capacity
Blots per run*
Primary antibodies
Valve delivered reagents
Recover primary reagents
Preprogrammed assays
User-defined assays
Steps per assay
Cycles per step
Rocking table
Volume delivery
Tubing
Alarm
Timer
Type of aspirate pump
Type of delivery pump
Pump speed (delivery)
Tray (Standard & Mini-blot)
Valves
Connectors
Power Supply requirement
Environmental requirements
Regulatory
Dimensions
Standard Tray
Mini-blot Tray
1
4
40 to 200 + ml
8–50 ml
1
4
One primary antibody fully automated. One primary
antibody, or four primaries semi-automated. The
Western Processor is programmed to alert the user
to manually add the different primary antibodies.
6 reagents
Yes. Second waste bottle or manual removal.
5
Up to 10
Up to 15
Up to 6
User selectable, 4 speed (0, slow, medium, fast)
0–500 ml
Medical grade Santoprene™, 1/8 inch (3 mm ID and
1 mm wall thickness) for waste: 1/16 inch (1.59 mm
ID and 0.8 mm wall thickness) for feed and valve
lines
Programmable alarm
0–999 minutes per step/cycle
DC diaphragm
Peristaltic
Two ml per second
High impact polystyrene (white)
Viton and U-polymer
Polypropylene
Globtek power supply, model#1240
• For indoor use only
• For use only up to 2000 meters
• To be operated only between 5 °C and 40 °C
• To be operated with a maximum relative
humidity of 80% for temperatures up to 31 °C,
decreasing linearly to 50% relative humidity at
40 °C
• Installation category I
• Pollution degree Z
CE, EN61010-1
26 (W) x 31 (D) x 13.7 (H) cm
Santoprene is a trademark of The Monsanto Company and exclusively licensed to Advanced Elastomer Systems, L.P.
* Two standard and eight mini blots may be processed at one time if placed in trays back to back. This may result in higher backgrounds
than if processed singly.
30
Appendix D
Worksheet for Programming a Method for the Western Processor
METHOD NAME:
(Up to 5 characters)
Valve # with pre-programmed names.
Valve 1: WASH1
Valve 2: WASH2
Valve 3: CONJ1
Valve 4: SUBST
(Valve 4 allows for a variable incubation time to monitor the development step)
Valve 5: CONJ2
Valve 6: REAGT
MANUL (Allows for manual addition of reagents, alarm will sound to prepare/add reagent)
Step #
Step # Reagent
Name/Valve #
Incubation
- hr. -min
Variable
Incubation
Valve4, subst only
Volume
-- ml
Not for Manul steps
Rocking
Speed?
Reagent
Alarm?
# of cycles
Max. 6 per step
Aspirate
after this
step?
Manul Aspirate
Volume
For Manul steps only
Step 1
Step 2
Step 3
Step 4
Step 5
Step 6
Step 7
Step 8
Step 9
Step 10
Step 11
Step 12
Step 13
Step 14
Step 15
Note: After running a method, it is very important to immediately wash all six (6) lines and valves using the "Purge Tubing Mode". If buffer salts and other reagents are allowed
to remain in the system, the lifespan of the valves will be greatly reduced.
Bio-Rad
Laboratories
Life Science
Group
2000 Alfred Nobel Drive
Hercules, California 94547
Telephone (510) 741-1000
Fax: (510) 741-5800
www.bio-rad.com
Bulletin 0000
US/EG
Rev A
Australia, Bio-Rad Laboratories Pty. Ltd., Block Y, Unit 1, Regents Park Industrial Estate, 391 Park Road, Regents Park, NSW 2143
Phone 02 9914 2800 • Fax 02 9914 2889
Austria, Bio-Rad Laboratories Ges.m.b.H., Auhofstraße 78D, A-1130 Wien •
• Fax (01)-876 56 29
•
•
• Fax 09-385 65 54
Brazil,
• Phone 55 21 507 6191
•
• Fax (905) 712-2990
China,
Phone 86-10-8201-1366/68 • Fax 86-10-8201-1367
Denmark,
•
• Fax 45 44 52-1001
•
• Fax 358 (0)9 804 1110
Finland, Bio-Rad Laboratories, Pihatörmö
France,
ö, 92430 Marnes-la-Coquette •
• Fax 01 47 41 9133
Germany, Bio-Rad Laboratories GmbH, Heidemannstraß
ö
önchen
Phone 089 318 84-177 • Fax 089 318 84-123
Hong Kong, Bio-Rad Pacific Ltd., Unit 1111, 11/F, New Kowloon Plaza, 38 Tai Kok Tsui Road, Tai Kok Tsui, Kowloon
Phone 852-2789-3300 • Fax 852-2789-1257
India, Bio-Rad Laboratories (India) Pvt. Ltd., B&B1, Enkay Towers Vanijyanikunj, Udhyog Vihar Phase V, Gurgaon, Haryana 122016
Phone (91-124)-6398112/113/114 • Fax (91-124)-6398115
Israel,
• Phone 03 951 4124 • Fax 03 951 4129
Italy, Bio-Rad Laboratories S.r.l., Via M. Peroglio 23, 00144 Rome • Phone 34 91 590 5200 •
Japan,
•
• Fax 03-5811-6272
•
•
Latin America,
•
• Fax 305-894-5960
•
The Netherlands,
•
Fax 0318-542216
•
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Norway,
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Russia,
•
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Singapore,
•
• Fax 65-2734835
Spain,
Sweden,
•
• Fax 46 (0)8-55 51 27 80
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United Kingdom,
• Free Phone 0800-181134 • Fax 01442-259118
00-000
0000
Sig 1200
4006145 Rev D
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