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Thermo
EASY-nLC 1200
User Guide for LC Devices
60053-97273 Revision A September 2015
© 2015 Thermo Fisher Scientific Inc. All rights reserved.
AFC, EASY-nLC, EASY-Column, EASY-Spray, Foundation, LC TriPlus, and Q Exactive are trademarks, and
Orbitrap, Orbitrap Fusion, Thermo Scientific, and Xcalibur are registered trademarks of Thermo Fisher
Scientific Inc. in the United States.
The following are registered trademarks in the United States: Advion and RePlay are registered trademarks of
Advion BioSystems, Inc. and Waters is a registered trademark of Waters Corporation.
The following are registered trademarks in the United States and other countries: Microsoft and Windows are registered trademarks of the Microsoft Corporation. McAfee and VirusScan are registered trademarks of
McAfee, Inc.
All other trademarks are the property of Thermo Fisher Scientific and its subsidiaries.
Thermo Fisher Scientific Inc. provides this document to its customers with a product purchase to use in the product operation. This document is copyright protected and any reproduction of the whole or any part of this document is strictly prohibited, except with the written authorization of Thermo Fisher Scientific Inc.
The contents of this document are subject to change without notice. All technical information in this document is for reference purposes only. System configurations and specifications in this document supersede all previous information received by the purchaser.
This document is not part of any sales contract between Thermo Fisher Scientific Inc. and a purchaser. This document shall in no way govern or modify any Terms and Conditions of Sale, which Terms and
Conditions of Sale shall govern all conflicting information between the two documents.
Release history: Revision A, September 2015
Software version: For the EASY-nLC device driver on LC Devices 3.0 or later, install Foundation platform version 3.0 SP2 or later and the Xcalibur data system version 3.0 or later.
For Research Use Only. Not for use in diagnostic procedures.
C
Contents
Thermo Scientific
Removing a Previous Version of the Instrument Control Software . . . . . . . . . . . 2
Installing the Current Version of the Instrument Control Software . . . . . . . . . . . 3
Excluding the EASY-nLC from the VirusScan Enterprise . . . . . . . . . . . . . . . . . . 4
Connecting the EASY-nLC Instrument to the Data System Computer . . . . . . . . 7
Adding the System Devices to the Instrument Configuration . . . . . . . . . . . . . . . 8
Setting Up the EASY-nLC Configuration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Setting Up the Sample Layout and Start Position . . . . . . . . . . . . . . . . . . . . . . . 15
Chapter 3 Modifying the HPLC Properties and Controlling Other Devices. . . . . . . . . . . . . .17
Accessing the Devices Page of the Touch-screen Application . . . . . . . . . . . . . . . 17
Adding the EASY-Spray Device to the Devices List . . . . . . . . . . . . . . . . . . . . . . 18
Setting Up the EASY-nLC Instrument Method Parameters. . . . . . . . . . . . . . . . 23
Sample Pickup and Loading Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
Pre-Column and Analytical Column Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
EASY-nLC 1200 User Guide for the Xcalibur Data System
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Contents
Setting Up the Injection Positions with the Sample Position Selector . . . . . . . . 43
Taking Corrective Action for Run-Time Errors. . . . . . . . . . . . . . . . . . . . . . . . . 48
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EASY-nLC 1200 User Guide for the Xcalibur Data System Thermo Scientific
P
Preface
This guide describes how to configure the Thermo EASY-nLC™ virtual instrument (VI) control software from the Thermo Foundation™ platform, set up the liquid chromatography
(LC) parameters for LC/MS instrument methods in the Instrument Setup window of the
Thermo Xcalibur™ data system, and set up the sample position settings and injection volumes for an injection sequence from the Sequence Setup view in the Xcalibur data system.
IMPORTANT To establish communication with the EASY-nLC instrument and to avoid the premature termination of data acquisition during sequence runs, set up the data system computer as described in
“Data System Computer Settings” on page viii
.
Contents
•
•
•
•
•
To suggest changes to the documentation or to the Help
Fill out a reader survey online at www.surveymonkey.com/s/PQM6P62 .
Thank you in advance for your help.
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Preface
Related Documentation
Thermo Fisher Scientific provides these manuals as PDF files on the data system computer and on the USB flash drive that ships with the EASY-nLC instrument:
• EASY-nLC 1200 Preinstallation Requirements Guide
• EASY-nLC 1200 Getting Started Guide (using the stand-alone, touch-screen application)
• EASY-nLC 1200 Troubleshooting and Maintenance Guide
• EASY-nLC 1200 User Guide for the Xcalibur Data System
The data system also provides Help for the EASY-nLC virtual instrument control software.
To access the EASY-nLC manuals on the data system computer
Choose Start > Programs (or All Programs) > Thermo Instruments > Manuals >
LC Devices > Thermo.
To open the Help from the EASY-nLC view of the Instrument Setup window
• To open the Welcome page of the Thermo EASY-nLC Help, choose Help > Thermo
EASY-nLC Help from the menu bar.
• To open the Help topic for the current page of the Thermo EASY-nLC view, do one of the following:
– Choose Help > Help On Current Item F1 from the menu bar.
– Press the F1 key on the computer keyboard.
To open the Help from the Thermo EASY-nLC Configuration dialog box of the
Foundation Instrument Configuration window
Press the F1 key on the computer keyboard.
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EASY-nLC Series User Guide for the Xcalibur Data System Thermo Scientific
Preface
System Requirements
Table 1 lists the data system computer hardware and software requirements for the EASY-nLC
VI control software (device driver). This device driver is provided on the LC Devices DVD and is designed for use with the Xcalibur data system or an equivalent Thermo Scientific™ mass spectrometry application.
IMPORTANT Before you install the device driver provided on the LC Devices DVD, ensure that the data system computer has a compatible version of the Foundation platform as noted in the Thermo LC Devices x.x.x Release Notes, where x.x.x is the version you want to install. You can find the Release Notes document, which includes the system requirements, on the LC Devices DVD.
Table 1. Data system computer hardware and software requirements
Hardware
Data system computer
Minimum requirements
• 2 GHz processor with 1 GB RAM
• DVD-ROM drive
• 80 GB hard drive capacity
• Display resolution 1280 × 1024
• 2 Network cards
• NTFS format
Requirements
Microsoft™ Windows™ 7 Professional (32-bit and 64-bit a
)
Software
Operating system
Microsoft .NET
Third-party software
Thermo Scientific software
4.5.1 for Foundation platform 3.1 or later
Adobe™ Reader™ 9.0 or later
LC Devices
EASY-nLC device driver
Foundation platform
3.0 or later 3.0 SP2 or 3.1
Touch-screen application For information about upgrading the touch-screen application for your EASY-nLC instrument, refer to the
EASY-nLC 1200 Troubleshooting and Maintenance Guide or the EASY-nLC 1200 Getting Started Guide.
a
LC Devices 2.8.0 and later are compatible with Windows 7, 32-bit and 64-bit
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Preface
Data System Computer Settings
Table 2 lists the computer settings that are required to avoid the premature termination of
data acquisition during sequence runs. It also provides the virus scanner setting that is required to establish communication with the EASY-nLC instrument.
IMPORTANT After you install the Thermo EASY-nLC device driver on a Windows 7 computer, exclude the Thermo EASY-nLC software from the Prevent IRC
Communication list for the computer’s virus scanner (see
“Excluding the EASY-nLC from the VirusScan Enterprise” on page 4
).
If you do not exclude the Thermo EASY-nLC software from the Prevent IRC
Communication list, the data system computer cannot establish communication with the
EASY-nLC instrument.
Table 2. Data system computer settings
Category
Power saver
Settings
• Screen saver: None
• Hibernate: Not enabled or Never
• Turn off hard disk: Never
Windows updates
Virus scanner
• Turn off monitor: Never
If the data system computer is connected to the Internet, turn off the automatic Windows updates.
For the Windows 7 operating system, exclude the
Thermo EASY-nLC software from the virus scanner.
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EASY-nLC Series User Guide for the Xcalibur Data System Thermo Scientific
Preface
Cautions and Special Notices
Make sure you follow the cautions and special notices presented in this guide. Cautions and special notices appear in boxes; those concerning safety or possible system damage also have corresponding caution symbols.
This guide uses the following types of cautions and special notices.
CAUTION Highlights hazards to humans, property, or the environment. Each CAUTION notice is accompanied by an appropriate CAUTION symbol.
IMPORTANT Highlights information necessary to prevent damage to software, loss of data, or invalid test results; or might contain information that is critical for optimal performance of the system.
Note Highlights information of general interest.
Tip Highlights helpful information that can make a task easier.
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Preface
Contacting Us
There are several ways to contact Thermo Fisher Scientific for the information you need. You can use your smartphone to scan a QR code, which opens your email application or browser.
Contact us Customer Service and Sales Technical Support
(U.S.) 1 (800) 532-4752 (U.S.) 1 (800) 532-4752
(U.S.) 1 (561) 688-8731 us.customer-support.analyze
@thermofisher.com
(U.S.) 1 (561) 688-8736 us.techsupport.analyze
@thermofisher.com
To find global contact information or customize your request
1. Go to www.thermoscientific.com
.
2. Click Contact Us, select the Using/Servicing a Product option, and then type the product name.
3. Use the phone number, email address, or online form.
To find product support, knowledge bases, and resources
Go to www.thermoscientific.com/support .
To find product information
Go to www.thermoscientific.com/lc-ms .
Note To provide feedback for this document:
• Send an email message to Technical Publications ( [email protected]
).
• Complete a survey at www.surveymonkey.com/s/PQM6P62 .
To order consumable and spare parts for the EASY-nLC instrument
For the EASY-nLC 1200 instrument, go to www.thermoscientific.com/nanoLCMS
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EASY-nLC Series User Guide for the Xcalibur Data System Thermo Scientific
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Installing the EASY-nLC Control Software
This chapter describes how to install the EASY-nLC virtual instrument (VI) control software from the LC Devices DVD. It also describes how to ensure communication between the data system computer and the EASY-nLC instrument.
Contents
•
Checking the System Prerequisites
•
Removing a Previous Version of the Instrument Control Software
•
Installing the Current Version of the Instrument Control Software
•
Excluding the EASY-nLC from the VirusScan Enterprise
Checking the System Prerequisites
The LC/MS system consists of the EASY-nLC instrument, a Thermo Scientific mass spectrometer, and a data system computer.
Before installing the EASY-nLC VI control software, do the following:
• Install compatible versions of the Foundation platform and the Xcalibur data system (or equivalent Thermo Scientific data system).
• Ensure that you have administrator rights to install the EASY-nLC VI control software for the Xcalibur data system (or equivalent Thermo Scientific data system).
• Remove any earlier version of the EASY-nLC VI control software from the data system
computer (see the next topic, Removing a Previous Version of the Instrument Control
).
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Installing the EASY-nLC Control Software
Removing a Previous Version of the Instrument Control Software
Before or after installing the EASY-nLC VI control software, do the following:
• Connect the EASY-nLC instrument to the data system computer through an Ethernet connection (see
“Connecting the EASY-nLC Instrument to the Data System Computer” on page 7
).
• Connect the EASY-nLC instrument to the Thermo Scientific MS detector with a contact closure cable.
The data system synchronizes the run timing between the LC system and the mass spectrometer through the cable. For information about connecting the contact closure cable, refer to the EASY-nLC 1200 Getting Started Guide.
• Check the laboratory local area network (LAN) port for Internet access to the remote support server. For remote diagnostics, the laboratory must have a LAN port with
Internet access.
The network firewall must allow outgoing Transmission Control Protocol/Internet
Protocol (TCP/IP) traffic from the EASY-nLC instrument to the remote support server at
IP address 46.226.219.10, port 22.
After installing the EASY-nLC VI control software, exclude the software from the virus scanner as described in
“Excluding the EASY-nLC from the VirusScan Enterprise” on page 4
.
Removing a Previous Version of the Instrument Control Software
If an older version of the instrument control software is installed on the data system computer, you must uninstall it from the Control Panel.
To remove a previously installed version of the EASY-nLC Xcalibur VI
1. From the Windows 7 desktop, choose Control Panel > Uninstall a Program.
The list of installed programs appears (see
).
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EASY-nLC 1200 User Guide for the Xcalibur Data System Thermo Scientific
1
Installing the EASY-nLC Control Software
Installing the Current Version of the Instrument Control Software
Figure 1.
Uninstall window for the Windows 7 operating system
2. Double-click Thermo LC Devices Thermo EASY-nLC Xcalibur VI and wait for the program to be removed from the list.
Installing the Current Version of the Instrument Control Software
The EASY-nLC VI control software provided with LC Devices 3.0 or later controls the
EASY-nLC instrument (with touch-screen software version 3.2 or later) and the column heater of the optional EASY-Spray ion source.
To install the EASY-nLC Virtual Instrument (VI) software
1. Check the installed versions of the Foundation platform and the Xcalibur data system and make sure that these versions are compatible with the LC Devices version that you are
installing (see “System Requirements” on page vii ).
2. Close all running programs before starting the installation.
3. If applicable, remove the previously installed version of the EASY-nLC VI control software (see
“Removing a Previous Version of the Instrument Control Software” on page 2
).
4. Load the LC Devices DVD into the data system computer, and follow the instructions in the Thermo LC Devices InstallShield Wizard.
5. On the Select Features page of the wizard, select the Thermo check box on the left, and
then select the Thermo EASY-nLC Xcalibur check box on the right (see Figure 2
).
Thermo Scientific EASY-nLC 1200 User Guide for the Xcalibur Data System
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1
Installing the EASY-nLC Control Software
Excluding the EASY-nLC from the VirusScan Enterprise
Figure 2.
Select Features page of the Thermo LC Devices InstallShield Wizard
6. When the installation finishes, restart the data system computer.
The installation is complete when you log back in.
Excluding the EASY-nLC from the VirusScan Enterprise
The McAfee™ VirusScan™ Enterprise software blocks the Thermo EASY-nLC software when the data system computer uses the Windows 7 operating system.
IMPORTANT For the Windows 7 operating system, you must exclude the EASY-nLC
VI software from the McAfee blocking list.
To exclude the EASY-nLC software from the VirusScan Enterprise 8.7.0i software
1. Log in to the data system computer as an administrator.
2. Open the McAfee VirusScan Console application as follows: a. Do one of the following:
• On the Windows 7 taskbar, right-click the McAfee icon, , and choose
Manage Features > VirusScan Enterprise.
–or–
• From the Windows 7 desktop, choose Start > All Programs > McAfee >
VirusScan Console.
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Installing the EASY-nLC Control Software
Excluding the EASY-nLC from the VirusScan Enterprise
3. In the Task column, right-click Access Protection and choose Properties from the shortcut menu.
The Access Protection Properties dialog box appears.
4. To open the Rules dialog box, do the following: a. Under Categories, choose Anti-virus Standard Protection.
b. Under Rules, select Prevent IRC Communication.
c.
Click Edit.
The Rules Details dialog box opens.
5. Modify these settings:
• In the Processes to Include box, delete the asterisk.
Note When the Processes to Include box contains only a single asterisk, the virus scanner prevents IRC communication for all processes.
• In the Processes to Exclude box, type Thermo*.
Note When the Processes to Exclude box contains the text Thermo*, the Access
Protection service for Internet Relay Chat (IRC) communication processes ignores processes that begin with Thermo.
6. Click OK to close the Rules Details dialog box.
7. Click OK to accept the new settings and close the Access Protection Properties dialog box.
8. Close the VirusScan Console application.
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2
Setting Up the Foundation Instrument Configuration
Connecting the EASY-nLC Instrument to the Data System Computer
Setting Up the Foundation Instrument Configuration
This chapter describes how to set up the instrument configuration for the EASY-nLC device driver.
Before you set up the instrument configuration, connect the EASY-nLC instrument to the data system computer, turn on the EASY-nLC instrument, and log in to the instrument as an administrator. Then, use the Thermo EASY-nLC Configuration dialog box of the Thermo
Foundation Instrument Configuration window to set up and test the network connection for the EASY-nLC instrument, to specify the tray type that you plan to use, and to identify the mobile phase solvents in solvent reservoir bottles A and B.
Contents
•
Connecting the EASY-nLC Instrument to the Data System Computer
•
Adding the System Devices to the Instrument Configuration
•
Setting Up the EASY-nLC Configuration
•
Setting Up the Sample Layout and Start Position
Connecting the EASY-nLC Instrument to the Data System Computer
The EASY-nLC instrument and the Thermo Scientific mass spectrometer communicate with the data system computer through an Ethernet connection routed through an Ethernet switch connected to the data system computer.
These are the default IP addresses of the data system computer and the EASY-nLC instrument.
Table 3. Default IP addresses for the LC system
System module
Data system computer
EASY-nLC instrument
IP address
172.16.0.101
172.16.0.103
To connect the EASY-nLC instrument to the data system computer
1. Connect a shielded Ethernet cable to the LAN port on the back panel of the EASY-nLC instrument.
2. Connect the other end of the Ethernet cable to an Ethernet switch.
3. Using a second shielded Ethernet cable, connect the Ethernet switch to the appropriate network card installed in the data system computer.
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Setting Up the Foundation Instrument Configuration
Adding the System Devices to the Instrument Configuration
For more information about setting up the communication cables, refer to the EASY-nLC
Series Getting Started Guide.
IMPORTANT Avoid IP conflicts with other units on the network, especially the mass spectrometer.
Adding the System Devices to the Instrument Configuration
Your Thermo Scientific LC/MS system consists of a Thermo Scientific mass spectrometer and an EASY-nLC instrument. To control these instruments from the Xcalibur data system, you must add the installed device drivers to the instrument configuration.
Note The Xcalibur data system and the Foundation Instrument Configuration window cannot be open simultaneously.
To add devices to the instrument configuration
1. Close the Xcalibur data system if it is open.
2. From the Windows taskbar, choose Start > Programs (or All Programs) >
Thermo Foundation
x.x > Instrument Configuration, where x.x is the installed version of the Foundation platform.
The Thermo Foundation Instrument Configuration window opens (see
). The installed devices are listed in the Available Devices list. The configured devices are listed in the Configured Devices list.
shows an instrument configuration with a mass spectrometer added to the
Configured Devices list.
Figure 3.
Thermo Foundation Instrument Configuration window
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Setting Up the Foundation Instrument Configuration
Setting Up the EASY-nLC Configuration
Tip To access Help, press F1 or click Help.
3. Add the EASY-nLC instrument to the instrument configuration as follows: a. Make sure that the Device Types list is set to All or AS (Autosampler).
If the Device Type list is set to LC, the EASY-nLC VI for Xcalibur device does not appear in the Available Devices list.
b. In the Available Devices list, double-click the Thermo EASY-nLC icon.
A copy of the Thermo EASY-nLC icon appears in the Configured Devices list.
4. Set up the configuration options for each configured device.
For information about the EASY-nLC configuration parameters, see the next topic,
“Setting Up the EASY-nLC Configuration.”
5. Click Done to close the Thermo Foundation Instrument Configuration window.
Tip You must close the Instrument Configuration window before you open the
Xcalibur data system. The two applications cannot be open at the same time.
Setting Up the EASY-nLC Configuration
Use the Thermo EASY-nLC Configuration dialog box to set up and test the communication connection to the data system computer, specify the autosampler model and the plate type that you are using, and identify the mobile phase solvents. Clicking Edit in the Autosampler
Configuration area (
) opens the Sample Position Selector dialog box where you set up the sample layout (row first or column first) and the start position for an Xcalibur injection sequence.
To set up the EASY-nLC configuration in the Foundation platform
1. If you have not already added the EASY-nLC instrument to the Configured Devices list of the Thermo Foundation Instrument Configuration window, add it as described in
“Adding the System Devices to the Instrument Configuration” on page 8 and leave the
Instrument Configuration window open.
2. In the Configured Devices list, double-click the Thermo EASY-nLC icon.
The Thermo EASY-nLC Configuration dialog box opens (see Figure 4 ).
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Setting Up the Foundation Instrument Configuration
Setting Up the EASY-nLC Configuration
Figure 4.
Thermo EASY-nLC Configuration dialog box
3. Ensure the following:
• The EASY-nLC instrument is connected to the data system computer by way of an
Ethernet switch (see
“To connect the EASY-nLC instrument to the data system computer” on page 7 ).
• The EASY-nLC instrument is turned on, and you are logged in as an administrator to the instrument.
4. Set up the network configuration for the EASY-nLC instrument as follows: a. Type the IP address of the EASY-nLC instrument and your user name in the appropriate boxes.
You must have an account in the EASY-nLC touch-screen application with the same user name. The default user name is “admin,” but you can change this to any existing user account on the EASY-nLC instrument.
Tip The EASY-nLC IP address is listed in the About box, which you can open by pressing the THERMO logo in the upper left corner of the touch screen. The current user is listed in the lower left corner of the touch screen.
If necessary, change the IP address on the Configuration > Network page of the touch-screen application. b. Click Test Connection to test the connection to the EASY-nLC instrument and to download the instrument configuration information to the data system computer.
5. Check the message in the status box to the right of the Test Connection button.
• If the connection is established, the status box changes to green (see
) and the information about the LC device driver and instrument appears.
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Setting Up the Foundation Instrument Configuration
Setting Up the EASY-nLC Configuration
• If a connection is not established, the status bar changes to red. Check the Ethernet cable connections, the IP address of the EASY-nLC instrument, and the IP address of the network card installed in the data system computer (see
EASY-nLC Instrument to the Data System Computer” on page 7 ).
• If an antivirus program or firewall blocks the connection, the application displays this error message:
An established connection was aborted by the software in your host machine.
Contact your local IT administrator for assistance.
For information about excluding the EASY-nLC software from the McAfee virus scanner, see
“Excluding the EASY-nLC from the VirusScan Enterprise” on page 4 .
6. Check the instrument information. If necessary, change the instrument information as described in
“Modifying the HPLC Properties” on page 20
.
7. Set up the autosampler configuration in the Autosampler Configuration area as follows: a. Select the autosampler model (Autosampler ASC).
b. Select the plate type (96- or 384-well plates, or a 6 × 8 (ASC) vial holder).
Note All plate formats for the ASC model have six extra vial positions on the side. When you set up an Xcalibur sequence, these vials are numbered 1-V1 to
1-V6.
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2
Setting Up the Foundation Instrument Configuration
Setting Up the EASY-nLC Configuration
Figure 5.
Thermo EASY-nLC Configuration dialog box
Touch-screen software version
4.0
Edit button c.
(Optional) Set up the default sample layout and start position to be used when
creating a new Xcalibur sequence as described in “Setting Up the Sample Layout and
8. In the Solvents for LC Pump area, type the names of the solvents in the boxes for solvent bottles A and B.
Solvent bottle A typically contains LC/MS-grade water with 0.1% formic acid. Solvent bottle B typically contains LC/MS-grade acetonitrile with 0.1% formic acid (max. 95% acetonitrile).
9. If the OK button is unavailable, click Test Connection to download the new configuration settings to the data system computer.
10. Click OK to accept the settings and close the Thermo EASY-nLC Configuration dialog box.
11. Click Done to close the Thermo Foundation Instrument Configuration window.
You must close the Foundation application before opening the Xcalibur data system. The
Xcalibur data system and the Thermo Foundation Instrument Configuration window cannot be open at the same time.
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Setting Up the Foundation Instrument Configuration
Setting Up the EASY-nLC Configuration
When you establish a connection between the EASY-nLC instrument and the data system computer, this dialog box displays the user-specified loop volume, idle flow rate, idle mixture, and column setup. You set these parameters from the EASY-nLC touch-screen application as
described above in “Modifying the HPLC Properties” on page 20
.
Table 4 describes the parameters in the Thermo EASY-nLC Configuration dialog box.
Table 4. EASY-nLC Configuration dialog box parameters (Sheet 1 of 2)
Parameter Description
EASY-nLC Network Configuration
Use this area to specify the IP address for the EASY-nLC instrument.
IP Address/Hostname Specifies the IP address of the EASY-nLC instrument. This value must match the value stored by the instrument.
Tip To check the IP address and host name stored by the instrument, press the company logo in the upper left corner of the EASY-nLC touch screen.
User Name Specifies the user name.
Note The user name that you type in this box must match the user name for the person currently logged in to the instrument. The user name is listed in the lower left corner of the EASY-nLC touch screen. For information about logging in to the EASY-nLC instrument, refer to the EASY-nLC Series Getting Started Guide.
Test Connection button
Tests the Ethernet connection between the data system computer and the EASY-nLC instrument. If the connection is established, the status box to the right of the button turns green and contains this message: Connection OK.
In addition, information for the LC device driver and instrument appears in the box below the status bar.
IMPORTANT If the following message appears, exclude the Thermo EASY-nLC
.
An established connection was aborted by the software in your host machine.
Contact your local IT administrator for assistance.
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2
Setting Up the Foundation Instrument Configuration
Setting Up the EASY-nLC Configuration
Table 4. EASY-nLC Configuration dialog box parameters (Sheet 2 of 2)
Parameter Description
LC Device Driver and Instrument Information
LC Device Driver
LC Instrument
Driver Info
Serial Number
Loop Volume
Idle Flow Rate
Displays the LC device driver version.
Displays the instrument type.
Displays the instrument’s touch-screen software version.
Displays the serial number that is listed on the instrument’s back panel.
Displays the user-specified loop volume stored by the EASY-nLC instrument.
Displays the user-specified idle flow rate that the instrument maintains at the end of an injection sequence.
Entering a value of 0 deactivates this feature.
Tip Maintaining long-term emitter stability (with glass emitters) might require a constant solvent flow through the system.
Idle Mixture
Column Setup
IFC
Displays the user-specified solvent composition (%B) that the instrument is to pump through the system at the end of an injection sequence.
Displays the user-specified selection. Displays One Column or Two Column.
Displays whether the intelligent flow control (IFC™) mode is enabled.
Max Pressure
Autosampler Configuration
Displays the maximum system pressure.
Use this area to select the autosampler type, plate type, sample layout, and initial vial position.
Autosampler Specifies the autosampler model:
Plate Installed
Sample Layout
Start Position
Edit button
• ASC—Autosampler with refrigerated compartment and 6 extra vials
Specifies the plate type that you plan to use: 96 well plate, 384 well plate, or 6 × 8 vial plate.
Displays the default sample layout (vial or microplate well positions) for an injection sequence.
Displays the default initial sample position for an injection sequence.
Opens the Sample Position Selector dialog box where you can select the sample layout and
initial position for an Xcalibur sequence (see “Setting Up the Sample Layout and Start
Solvents for LC Pump
Use this area to describe the solvents that make up the gradient.
A
B
Specifies the solvent in solvent bottle A.
Specifies the solvent in solvent bottle B.
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Setting Up the Foundation Instrument Configuration
Setting Up the Sample Layout and Start Position
Setting Up the Sample Layout and Start Position
When you open the Sample Position Selector dialog box from the Thermo EASY-nLC
Configuration dialog box, the Number of Samples to Select box and the Select button are unavailable.
When setting up the configuration options for the EASY-nLC instrument, use the Sample
Position Selector dialog box to set up the sample layout and initial vial or microplate well position for an injection sequence.
For information about using the Sample Position Selector dialog box to interactively select the
vial or microplate well positions for an injection sequence, see “Setting Up the Injection
Positions with the Sample Position Selector” on page 43
.
To set up the sample layout and start position for an Xcalibur injection sequence
1. In the Autosampler Configuration area, click Edit.
The Thermo EASY-nLC Sample Position Selector dialog box opens. The Number of
Samples to Select box is unavailable (see Figure 6 ).
Figure 6.
Sample Position Selector dialog box
Thermo Scientific
2. In the Sample Positions area, click Clear All.
3. In the Plate Rack area, select the default initial sample position by clicking a position.
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Setting Up the Foundation Instrument Configuration
Setting Up the Sample Layout and Start Position
4. In the Sample Selection Layout area, select the Row First or Column First option.
5. Click OK to save your changes and close the Thermo EASY-nLC Sample Position
Selector dialog box.
Note You can also open the Thermo EASY-nLC Sample Position Selector dialog box by clicking Select Vials in the Samples area of the Xcalibur New Sequence Template
dialog box (see Figure 16 on page 40
).
Table 5 describes the parameters in the Sample Position Selector dialog box. When you access
this dialog box from the EASY-nLC Configuration dialog box, only the Sample Position
Layout and Initial Position parameters are available.
Table 5. Sample Position Selector dialog box parameters
Parameter
Plate Rack
Description
Use this area to select or clear the sample positions.
Vial Rack
For an EASY-nLC instrument with an ASC model autosampler, you can select or clear a sample position from the additional 6 vials.
Note The additional vials are located on rack 1 and are called 1-V1 to 1-V6.
Sample Selection Layout
Row First
Column First
When you select the Row First option, the data system fills the sequence table with the sample positions row by row.
When you select the Column First option, the data system fills the sequence table with the sample positions column by column.
Note The sequence editor ignores the Sample Selection Layout option when you select the vial or well positions by clicking a set of samples in the Plate and Vial Rack areas.
Sample Positions (click on plate)
Initial Position
Number of Samples to
Select
Clear All button
Select button
Specifies the first vial or microplate well position in the sequence table.
Specifies the number of samples to add to the sequence table in addition to the initial vial position.
Clears all of the Plate and Vial Rack entries and the Initial Position entry.
Sequentially selects the number of vial or well positions specified in the Number of Samples to Select box. The selected vials or wells are highlighted in green in the Plate Rack and Vial
Rack areas. The vials or wells are ordered sequentially according to the sample selection layout selection.
Dialog box buttons
OK
Cancel
Accepts the settings and closes the dialog box.
Closes the dialog box without accepting the changes.
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Modifying the HPLC Properties and Controlling
Other Devices
This chapter describes how to set up the HPLC properties for the EASY-nLC instrument and how to set up control for the EASY-Spray device from the EASY-nLC instrument’s touch-screen application.
Before you can control an external device, such as the EASY-Spray ion source’s column heater, you must add the device to the Devices list on the Maintenance > Devices page of the touch-screen application. You also set up the HPLC properties for the EASY-nLC instrument on the Maintenance > Devices page.
Contents
•
Accessing the Devices Page of the Touch-screen Application
•
Adding the EASY-Spray Device to the Devices List
•
Accessing the Devices Page of the Touch-screen Application
Use the Maintenance > Devices page of the touch-screen application to modify the
EASY-nLC instrument’s HPLC properties and the add the EASY-Spray ion source’s column heater to the list of devices that the EASY-nLC instrument can control.
To access the Maintenance > Devices page
1. Log in to the EASY-nLC instrument as an administrator. For information about logging into the instrument, refer to the EASY-nLC Series Getting Started Guide.
When you log in as an administrator, all five menu tabs on the menu bar at the top of the screen.
2. From the menu bar at the top of the screen, press Maintenance.
The vertical tabs on the far left of the screen are Scripts, Log Book, Support, and Devices.
3. In the vertical tabs, press Devices.
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Modifying the HPLC Properties and Controlling Other Devices
Adding the EASY-Spray Device to the Devices List
The Maintenance > Devices page opens (
Figure 7.
Maintenance Devices page
Adding the EASY-Spray Device to the Devices List
The EASY-nLC instrument can control the EASY-Spray ion source’s column temperature.
Note The EASY-Spray ion source connects to the mass spectrometer’s ion source interface. The EASY-nLC instrument provides power for the ion source through a P-Bus connection. For information about setting up the EASY-Spray ion source, refer to the
EASY-Spray Series Ion Source User Guide. For information about connecting the P-Bus cable to the EASY-nLC instrument, refer to the EASY-nLC 1200 Getting Started Guide.
To add the EASY-Spray ion source to the Devices list
1. On the instrument’s touch screen, press Maintenance > Devices and check that the
EASY-Spray device is not already present in the Devices list. For more information about accessing the Maintenance Devices page, see
“Accessing the Devices Page of the
Touch-screen Application” on page 17 .
2. To add the EASY-Spray device, do the following: a. Press Add Device.
The Select a Device to Add dialog box opens ( Figure 8 ).
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Modifying the HPLC Properties and Controlling Other Devices
Adding the EASY-Spray Device to the Devices List
Figure 8.
Select a Device to Add dialog box b. Select EASY-Spray and press Accept.
3. Verify that the Devices list includes the added device.
Selecting the device displays information about the device driver on the About page
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Modifying the HPLC Properties and Controlling Other Devices
Modifying the HPLC Properties
Figure 9.
About information for the EASY-Spray device
Modifying the HPLC Properties
Use the Properties page for the EASY-nLC HPLC device to do the following:
• Enter the loop volume of the sample loop that you are using in the EASY-nLC instrument.
• Set up the solvent system to deliver a low flow of solvent when the instrument is idle.
• Specify the column setup that you are using.
• Turn on the Load Speed Protection feature.
• Turn on the Automatic Pre-run Maintenance feature.
To modify the device properties
1. From the instrument’s touch screen, do the following: a. Open the Maintenance > Devices page.
b. In the Devices list, select EASY-nLC (HPLC), and then press the Properties tab.
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Modifying the HPLC Properties and Controlling Other Devices
Modifying the HPLC Properties
The Properties view for the HPLC device on the Maintenance > Devices page opens
).
Figure 10. EASY-nLC (HPLC) Properties view
Thermo Scientific c.
Press the corresponding box to open the keypad and enter the appropriate numeric values for these parameters:
• For Loop Volume, type the nominal volume of the sample loop installed on valve S (5, 20, or 50 μL).
• For Idle Flow Rate, type an appropriate value, in nL/min. When idle, the instrument pumps the specified solvent mixture at the specified flow rate.
• For Idle Mixture %B, type a value from 0 to 100.
d. Select or clear these check boxes as appropriate: One Column Setup, Load Speed
Protection (LSP), and Automatic Pre-run Maintenance.
Note These check boxes are available in the following versions of the EASY-nLC touch-screen application:
• One Column Setup
• LSP
• Automatic Pre-run Maintenance e.
Click Apply.
2. In the EASY-nLC Configuration dialog box of the Foundation Instrument Configuration window, click Test Connection. Then verify the new settings.
For information about the EASY-nLC Configuration dialog box, see
EASY-nLC Configuration” on page 9
.
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Creating Xcalibur Instrument Methods
This chapter describes how to set up the instrument method parameters for the EASY-nLC instrument.
An Xcalibur instrument method (.meth) contains the chromatographic and data acquisition parameters for your LC/MS system. When creating new instrument methods, set up the parameters for all of the configured devices.
Contents
•
Setting Up the EASY-nLC Instrument Method Parameters
•
Note For information about setting up the instrument method parameters for the mass spectrometer, refer to the Help for the mass spectrometer.
Setting Up the EASY-nLC Instrument Method Parameters
Use the Instrument Setup window of the Xcalibur data system to create instrument methods for your LC/MS system. To create an instrument method, enter the data acquisition settings for the mass spectrometer and the injection and chromatographic settings for the LC system.
To open the Instrument Setup window
From the Xcalibur Home Page, do one of the following:
• Choose GoTo > Instrument Setup.
–or–
• Click the Instrument Setup icon in the Roadmap view of the Home Page.
The Instrument Setup window opens. The view bar contains icons for the configured devices.
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
To set up the instrument method parameters for the EASY-nLC instrument
1. In the view bar of the Instrument Setup window, click the Thermo EASY-nLC icon.
The EASY-nLC view opens in the Instrument Setup window with the Sample Pickup and
Loading page displayed (see Figure 11
). At the bottom of the view, the Solvents area lists the solvents specified in the instrument configuration. To change the names of the mobile
phase solvents, see “Setting Up the EASY-nLC Configuration” on page 9
.
2. Enter the appropriate instrument method settings for the EASY-nLC instrument. Click
Next to open the remaining three pages of the EASY-nLC view page by page.
Figure 11 shows the Sample Pickup and Loading page of the EASY-nLC view.
Figure 11. EASY-nLC view of the Instrument Setup window
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
For information about the pages of the EASY-nLC view, see these topics:
•
Sample Pickup and Loading Page
next topic
•
•
“Pre-Column and Analytical Column Page” on page 30
•
•
Sample Pickup and Loading Page
Use the Sample Pickup and Loading page to specify the following:
• The sample volume that the autosampler withdraws (injection volume) from the vial or well position in the Xcalibur injection sequence and the flow rate at which the autosampler draws the sample into the sample loop
• The solvent volume that pump A uses to push the sample out of the sample loop and onto the column assembly and the flow rate and maximum pressure for the sample loading step
To open the Sample Pickup and Loading page
1. Open the EASY-nLC view of the Instrument Setup window.
2. If the Sample Pickup and Loading page is not displayed, click the Sample Pickup and
Loading tab.
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
Table 6 describes the parameters on the Sample Pickup and Loading page of the EASY-nLC
view of the Instrument Setup window.
Table 6. Sample Pickup and Loading page parameters
Parameter
Sample Pickup
Volume (μL)
Description
Flow (μL/min)
Specifies the sample volume that the autosampler withdraws from the vial or well position specified in the Xcalibur sequence.
Maximum volume = sample loop volume – 2 μL
The graphic on the Home > Overview page (touch screen) displays the sample loop size.
Specifies the flow rate at which pump S draws the sample into the sample loop.
Range: 0 to 40 μL/min (The flow must be greater than 0 when the sample pickup volume is greater than 0.)
Tip For aqueous sample matrixes, set the flow rate to 20 μL/min.
Sample Loading
Volume (μL) Specifies the volume of solvent A that the instrument uses to push the sample out of the sample loop and onto the precolumn (two-column setup) or the analytical column
(one-column setup).
Typically, two times the sample pickup volume plus 2 μL is sufficient.
Range: 0 to 137 μL
IMPORTANT The maximum allowable entry is 137 μL; however, for Xcalibur sequence runs, the sample loading volume plus the precolumn equilibration volume plus the analytical
column equilibration volume must be less than 120 μL.
Flow (μL/min)
See “Taking Corrective Action for Run-Time Errors” on page 48 .
Specifies the flow rate at which pump A pumps solvent A through the instrument to push the sample out of the loop and onto the precolumn or analytical column.
If the Flow box is empty, the instrument runs at the set pressure.
Range: 0 to 100 μL/min
Specifies the maximum pressure for the sample loading step. Max Pressure (bar)
If the Max Pressure box is empty, the instrument runs at the set flow rate.
EASY-nLC 1200 range: 0 to 1180 bar
Tip If both the Max Pressure and Flow boxes are empty, the instrument runs at the maximum instrument pressure.
Leave these boxes empty if you want the instrument to run at the highest possible flow rate that is allowed under the maximum instrument pressure limit.
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
Gradient Page
Use the Gradient page to set up the gradient program (see
To open the Gradient page
1. Open the EASY-nLC view of the Instrument Setup window.
2. Click the Gradient tab. If you do not see the Gradient tab, click Next.
Figure 12. Gradient page
Thermo Scientific
To set up the gradient program
1. In the Time = 0.00 row, do the following:
• In the Flow (nL/min) column, type a value from 1 to 2000.
• In the %B column, type a value from 0 to 100.
2. To add an additional time point (table row), click Add.
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
3. In rows 2 or higher of the gradient table, do the following: a. In the Duration column, type a value from 0.01 to 9999.49.
The value in the Duration column is the length of time that you want the instrument to take to reach the specified flow and %B conditions. When you type a value in the
Duration column, the value in the Time column changes to the current Duration value plus the value in the previous row. b. In the Flow (nL/min) column, type a value from 1 to 2000.
c.
In the %B column, type a value from 0 to 100.
Both the flow rate and %B composition change linearly between time points.
4. Edit the table as follows:
• To delete a row, select the row and click Del.
• To reorder the rows as applicable, select the row and click Up or Down.
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Setting Up the EASY-nLC Instrument Method Parameters
Table 7 describes the parameters on the Gradient page.
Table 7. Gradient page parameters
Parameter
Table
Time
Description
Duration
Specifies the run time when the instrument reaches the conditions specified in the flow and %B columns.
Specifies the length of time between the previous and current time points.
Flow (nL/min)
Range: 0.01 to 9999.59 min
Specifies the mobile phase flow rate for the current time point.
%B
Range: 1 to 2000 nL/min
Specifies the percent composition of solvent B in the mobile phase for the current time point.
Buttons
Add
Del
Up
Down
Graph
Flow (nL/min)
Adds a row to the gradient program.
Deletes the selected row from the gradient program.
Moves the selected row up.
Moves the selected row down.
AB mixture (%B)
Time (min)
The y axis on the left side of the graph displays the flow scale.
A black line represents the change in flow rate with time.
The y axis on the right side of the graph displays the percentage of solvent B.
A red line represents change in %B with time.
The x axis displays the run time of the gradient program.
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
Pre-Column and Analytical Column Page
Use the Pre-column and Analytical Column page to set up the column equilibration steps
(see
To open the Pre-column and Analytical Column page
1. Open the EASY-nLC view of the Instrument Setup window.
2. Click the Pre-column and Analytical Column tab. If you do not see the Pre-column and
Analytical Column tab, click Next.
Figure 13. Pre-column and Analytical Column page
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Setting Up the EASY-nLC Instrument Method Parameters
Table 8 describes the parameters on the Pre-column and Analytical Column page.
Note For information about determining the appropriate flow rate for the precolumn and analytical column equilibration steps, refer to the EASY-nLC 1200 Getting Started Guide.
Table 8. Pre-column and Analytical Column page parameters (Sheet 1 of 2)
Parameter
Pre-column Equilibration
Description
Volume (μL) Specifies the volume of solvent A that pump A pushes through the precolumn to waste during the precolumn equilibration step.
Flow (μL/min)
Max Pressure (bar)
Range: 0 to 137 μL
IMPORTANT The maximum allowable entry is 137 μL; however, for Xcalibur sequence runs, the sample loading volume plus the precolumn equilibration volume plus the analytical
column equilibration volume must be less than 120 μL.
Specifies the flow rate for the precolumn equilibration step. If the Flow box is empty, the instrument runs at the set pressure.
Range: 0 to 100 μL/min
Specifies the maximum pressure for the precolumn equilibration step. If the Max Pressure box is empty, the instrument runs at the set flow rate.
EASY-nLC 1200 range: 0 to 1180 bar
Tip If both the Max Pressure and Flow boxes are empty, the instrument runs at the maximum instrument pressure.
Leave these boxes empty if you want the instrument to run at the highest possible flow rate that is allowed under the maximum instrument pressure limit.
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
Table 8. Pre-column and Analytical Column page parameters (Sheet 2 of 2)
Parameter Description
Analytical Column Equilibration
Volume (μL) Specifies the volume of solvent A that pump A pushes through the analytical column during the analytical column equilibration step.
Flow (μL/min)
Range: 0 to 137 μL
IMPORTANT The maximum allowable entry is 137 μL; however, for Xcalibur sequence runs, the sample loading volume plus the precolumn equilibration volume plus the analytical
column equilibration volume must be less than 120 μL.
See “Taking Corrective Action for Run-Time Errors” on page 48 .
Specifies the flow rate for the analytical column equilibration step. If the Flow box is empty, the instrument runs at the set pressure.
Range: 0 to 3 μL/min
Specifies the maximum pressure for the analytical column equilibration step.
Max Pressure (bar)
EASY-nLC 1200 range: 0 to 1180 bar
Note If both the Max Pressure and Flow boxes are empty, the instrument runs at the maximum instrument pressure. Leave these boxes empty if you want the instrument to run at the highest possible flow rate that is allowed under the maximum instrument pressure limit.
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Setting Up the EASY-nLC Instrument Method Parameters
Autosampler Page
Use the Autosampler page to set up the wash step for the needle and sample loop ( Figure 14 ).
To open the Autosampler page
1. Open the EASY-nLC view of the Instrument Setup window.
2. Click the Autosampler tab. If you do not see the Autosampler tab, click Next until the tab appears.
Figure 14. Autosampler page
Thermo Scientific
To set up the standard autosampler wash step
1. Select the Standard option.
2. In the Flush Volume (μL) box, type the volume of solvent A (from wash bottle 3) that the autosampler uses to flush the needle and the sample loop.
The range depends on the sample loop size (see
).
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Creating Xcalibur Instrument Methods
Setting Up the EASY-nLC Instrument Method Parameters
To set up a custom wash program
1. Select the Custom option.
The custom wash table becomes available.
2. For each row that you want to add to the program, click Add.
You can select a different wash solvent for each row.
3. Edit each row as follows:
• In the Source column, select the wash solvent source: Bottle 1, Bottle 2, or Bottle 3.
IMPORTANT The EASY-nLC tray compartment holds up to four wash bottles:
• Bottles 1 and 2 in positions W1 and W2 can contain custom wash solvents.
• Bottle 3 in position W3 must contain solvent A.
• Bottle 4 in position W4 must contain the needle wash insert.
During a custom wash program, the autosampler does the following:
1. Empties the contents of pump S into the needle wash bottle (W4).
2. Performs the steps in the custom wash program. During each cycle, the autosampler draws solvent from the specified bottle and then ejects the solvent into the needle wash bottle (W4).
3. Performs a final wash cycle with a preset volume of solvent A from bottle 3
(W3).
4. Refills pump S with solvent A from bottle 3 (W3) to be ready for the next injection.
• In the Volume (μL) column, type a volume from 0 to the sample loop size plus 8 μL.
If you enter a volume that is higher than 58 μL, an Invalid Method Parameter message box appears and the program resets the volume to the maximum allowed.
• In the Cycles column, type a value from 0 to 10.
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Setting Up the EASY-nLC Instrument Method Parameters
Table 9 describes the parameters on the Autosampler page.
Table 9. Autosampler page parameters (Sheet 1 of 2)
Parameter
Standard
Standard option
Flush Volume (μL)
Description
Selecting this option enables the standard wash option.
Specifies the flush volume that pump S is to push through the sample loop.
Range for the 20 μL sample loop: 30.25 to 100.75 μL
Range for the 50 μL sample loop: 60.26 to 70.75 μL
Custom
Custom option Selecting this option enables the custom wash program.
For information about the steps that the autosampler performs during a custom wash program, see
“To set up a custom wash program” on page 34 .
Custom wash table columns
Order
Source
Defines the order of the custom wash steps.
Specifies the wash solvent source for the current wash step.
Volume (μL)
Selections: Bottle 1, Bottle 2, or Bottle 3
Note Bottles 1 and 2 contain custom wash solvents. Bottle 3 contains solvent A.
Specifies the wash solvent volume for each wash cycle.
Maximum entry volume: 58 μL
Cycles
Maximum allowed volume at run time: Sample loop size plus 8 μL
Note The method editor of the EASY-nLC VI control software does not know the configured sample loop size. When you submit a sequence with the instrument method, the data system’s acquisition service validates the custom wash volumes against the configured sample loop size. To prevent solvent from being drawn into pump S, the instrument control software limits the volume to the sample loop size plus 8 μL.
For information about changing the sample loop size on the Maintenance > Devices page of the touch-screen application, see
“Modifying the HPLC Properties” on page 20 .
Specifies the number of wash cycles for the wash step.
Range: 1 to 10
Note For each cycle, the autosampler draws the specified volume of wash solvent from the specified source into the needle and then ejects the wash solvent into wash bottle 4 (W4).
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Setting Up the EASY-nLC Instrument Method Parameters
Table 9. Autosampler page parameters (Sheet 2 of 2)
Parameter
Buttons
Add
Del
Up
Down
Description
Adds a row to the wash program.
Deletes the selected row from the wash program.
Moves the selected row up.
Moves the selected row down.
EASY-Spray Page
Use the EASY-Spray page to control the temperature of the column heater during a sequence run.
To open the EASY-Spray page
1. Open the EASY-nLC view of the Instrument Setup window.
2. Click the EASY-Spray tab. If you do not see the EASY-Spray tab, click Next until the tab appears (
Tip If the Autosampler tab is the last tab, check whether the Device list on the
Maintenance > Devices page of the touch-screen application lists the EASY-Spray
column heater (see “Accessing the Devices Page of the Touch-screen Application” on page 17
).
Figure 15. EASY-Spray page with the default settings
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Setting Up the EASY-nLC Instrument Method Parameters
To specify the column heater temperature
1. On the EASY-Spray page, select the Column Heating check box.
2. Type a temperature from 30 to 60 °C.
Tip If you cannot control the column heater temperature, check the following:
• If the temperature readout on the EASY-Spray ion source is blank, make sure that the
P-Bus cable is properly connected and that the EASY-nLC instrument is turned on.
• If the temperature readout in the EASY-Spray dialog box (available from the Home >
Overview page of the touch-screen application) displays a question mark and the temperature readout on the EASY-Spray ion source displays two vertical bars, make sure that the EASY-Spray column is plugged into the EASY-Spray ion source’s Heater port.
For more information about the EASY-Spray ion source, refer to the EASY-Spray Series Ion
Source User Guide.
Table 9 describes the parameters on the EASY-Spray page.
Table 10. EASY-Spray page parameters
Parameter
Column Heating
Description
Selecting this check box turns on the remote column heater control. Clearing this check box turns off the remote column heater control.
Column Temperature
(T)
Default: Clear
Specifies the temperature setting for the EASY-Spray column. The data system downloads this setting to the EASY-Spray ion source at the beginning of a sample run.
Default: 30 °C
Range: 30–60 °C
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Creating Xcalibur Instrument Methods
Saving Instrument Methods
Saving Instrument Methods
After you set up the instrument method parameters for the LC/MS system, save the instrument method to an appropriate directory.
To save your current instrument method
1. Choose File > Save As.
The Save As dialog box opens.
2. In the File Name box, type a name for the new instrument method.
3. Click Save.
The File Summary Information dialog box opens.
4. Click OK.
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Creating and Running Sequences
Creating Sequences
Creating and Running Sequences
This chapter describes how to create and run sequences in the Xcalibur Sequence Setup view.
Contents
•
•
Setting Up the Injection Positions with the Sample Position Selector
•
•
Taking Corrective Action for Run-Time Errors
Creating Sequences
In the Xcalibur data system, the table that contains the sample injection information is called a sequence. In addition to the data file name for each injection, the sequence must contain the vial or microplate well positions of the samples (unknowns, calibration standards, quality control check standards, and so on) in the sample set, the instrument method to be used, and the injection volume. You can specify a different instrument method for each injection.
Tip You cannot change the tray type for the EASY-nLC instrument in the Sequence Setup view of the Xcalibur Home Page. If you choose Change > Tray Name from the menu bar, the following Home Page message appears:
The configured autosampler does not offer any selection of trays.
To change the tray type, close the Xcalibur data system. Then, open the EASY-nLC
Configuration dialog box as described in “Setting Up the EASY-nLC Configuration” on page 9
and select the appropriate tray type.
To create sequences in the Xcalibur data system
1. Do one of the following to open the Home Page – Sequence Setup view:
• From the Home Page – Roadmap view, click the Sequence Setup icon, .
• From any Home Page view, choose View > Sequence Setup View.
• From the Instrument Setup window, click the Home Page icon, , and then choose View > Sequence Setup View.
2. To open the New Sequence Template dialog box, choose File > New.
The New Sequence Template dialog box opens (see
).
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Creating and Running Sequences
Creating Sequences
Figure 16. New Sequence Template dialog box
For the EASY-nLC instrument, you cannot change the tray type in the Sequence Setup view. Tray Type is a read-only parameter.
3. In the General area, do the following:
• Type the base file name for the data files.
• Type a starting number for the value that the data system appends to the base file name.
• Browse to select the directory location for the data files in the Path box.
• Browse to select the instrument method in the Instrument Method box.
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5
Creating and Running Sequences
Creating Sequences
4. Set up the vial or well positions by doing one of the following:
• To use the text boxes in the Samples area of the New Sequence Template dialog box, go to
.
–or–
• To use the Sample Position Selector dialog box, go to “Setting Up the Injection
Positions with the Sample Position Selector” on page 43
.
Tip You can set up the sample positions for a sequence in three ways:
• In the Samples area of the New Template Sequence dialog box, type the number of samples in the Number of Samples box and the starting vial position in the Initial Vial Position box.
• In the Sample Position Selector dialog box, do one of the following:
– Type the number of samples in the Number of Samples to Select box, and then click Select (see
).
– Click the positions in the Plate Rack and Vial Rack areas. Use this method to set up noncontiguous, nonsequential, or noncontiguous and nonsequential positions.
5. Set up the vial or well positions by using the Samples area of the New Sequence Template dialog box as follows:
• In the Number of Samples box, type the number of samples for the injection sequence.
• In the Injections per Sample box, type the number of injections that you want the autosampler to make for each sample.
• For replicate sample injections, do the following:
– Select the Re-Use Vial Positions check box if you want the autosampler to make replicate injections from the same sample vial or well. Make sure that the vial or well contains a sufficient volume.
– Clear the Re-Use Vial Positions check box if you want the autosampler to make replicate injections from different sample vials or wells. Load the autosampler with a sufficient number of replicate samples.
• In the Initial Vial Position box, type the vial position for the first sample injection.
For alphanumeric entries, use uppercase letters (see
).
EASY-nLC 1200 User Guide for the Xcalibur Data System
41
5
Creating and Running Sequences
Creating Sequences
Note The sequence editor fills in the Positions column of the sequence table with contiguous vial or well locations based on the initial position, the number of samples, the number of replicate injections, and the sample selection layout option selected in the Foundation Instrument Configuration window.
• Selecting the Row First option in the instrument configuration application prompts the sequence editor to fill in the positions row by row (for example,
A1–A8, B1–B8, and so on).
• Selecting the Column First option in the instrument configuration application prompts the sequence editor to fill in the positions column by column (for example, A1–F1, A2–B2, and so on).
• Clearing the Re-Use Vials check box prompts the sequence editor to enter an additional vial or well position for each replicate.
6. To set up the remaining parameters in the New Sequence Template dialog box, refer to the Sequence Setup view Help.
7. Click OK to create the sample sequence.
The sequence information appears in the Sequence Setup view (see
Figure 17. Sequence Setup view on the Xcalibur Home Page
When the Inj Vol column lists a value of –1, the instrument uses the Sample Pickup volume that is specified in the instrument method.
8. Check the following:
• The vial or well values in the Position column—Make sure that the entries are valid and use uppercase letters.
• The numeric value in the Inj Vol column—When the value is –1, the data system uses the sample pickup volume specified in the instrument method.
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Creating and Running Sequences
Setting Up the Injection Positions with the Sample Position Selector
IMPORTANT The sequence table contains a column called Inj Vol (injection volume). If the column does not appear, choose Change > Column
Arrangement and add the Inj Vol column to the displayed columns.
To ensure that the injection volume specified in the instrument method file is used as intended, the default value in the Inj Vol column is set to –1, which is an invalid injection volume. When the Xcalibur data system detects the invalid value during a run, it automatically takes the actual injection volume from the corresponding instrument method file, overriding the –1 with the correct value.
During a run, the autosampler draws the specified injection volume (sample pickup volume) into the sample loop. The system then flushes the sample out of the sample loop and onto the LC column with the specified sample loading volume. Specifying an insufficient loading volume can result in blank injections or poor injection reproducibility because the system is either not loading or only partially loading the sample onto the LC column.
Changing the sample pickup volume by specifying a different value in the Inj Vol column of the sequence does not change the sample loading volume specified in the instrument method. To prevent incompatible settings, avoid using the Inj Vol column to vary the injection volume. Instead, create different instrument methods, where each instrument method contains compatible settings for the sample loading and sample pickup volumes (see
). You can use a different instrument method for each sequence row.
Setting Up the Injection Positions with the Sample Position Selector
For information about the parameters in the Sample Position Selector dialog box, see Table 5
on
. The functionality of the Sample Position Selector dialog box depends on where you access the dialog box:
• When you open the Sample Position Selector dialog box from the EASY-nLC
Configuration dialog box of the Thermo Foundation > Instrument Configuration window, use the dialog box to set up the sample layout and initial vial or microplate well
position for an injection sequence (see “Setting Up the Sample Layout and Start Position” on page 15
).
• When you open the Sample Position Selector dialog box from the New Sequence
Template dialog box in the Xcalibur Sequence Setup view, use the dialog box to interactively set up the vial or microplate well positions for the injection sequence as described in the following procedure.
Thermo Scientific EASY-nLC 1200 User Guide for the Xcalibur Data System
43
5
Creating and Running Sequences
Setting Up the Injection Positions with the Sample Position Selector
Note The sample position nomenclature is as follows:
• For 6 × 8 vials, the rows are labeled A–F and the columns are labeled 1–8. The nomenclature for the column A and row 1 position is A1.
• For the large vials in the Vial Rack area, the nomenclature is 1-V1 to 1-V6.
• For the microplate wells of a 96-well microplate, the nomenclature for the well positions is A1 to H12.
• For the microplate wells of a 384-well microplate, the nomenclature for the well positions is A1 to P24.
To open the Sample Position Selector dialog box from the New Sequence Template dialog box and set up the sample positions
1. Click Select Vials in the Samples Area (see Figure 16 on page 40
).
The Thermo EASY-nLC Sample Position Selector dialog box opens (see
). The rows for the 6 × 8 vial tray are labeled A–F, and the columns are labeled 1–8. The six additional vials in the Vial Rack area are numbered 1-V1 to 1-V6.
Figure 18. Thermo EASY-nLC Sample Position Selector dialog box
2. Follow one of these procedures:
•
To select vials or wells by using the Plate Rack and Vial Rack areas
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EASY-nLC 1200 User Guide for the Xcalibur Data System Thermo Scientific
Thermo Scientific
5
Creating and Running Sequences
Setting Up the Injection Positions with the Sample Position Selector
•
To select contiguous vials or wells in sequential order by using the Select button
3. To set up the remaining parameters in the New Sequence Template dialog box, refer to the Sequence Setup view Help.
4. Click OK to create the sample sequence.
The sequence information appears in the Sequence Setup view on the Xcalibur Home
Page (see Figure 17 on page 42 ).
To select vials or wells by using the Plate Rack and Vial Rack areas
1. In the Sample Positions (Click on Plate or Select Button) area, click Clear All.
This clears the Initial Position box and any selected positions on the Plate Rack and the
Vial Rack.
2. Select contiguous or noncontiguous vials or wells in a sequential or nonsequential order by clicking the positions on the Plate Rack and Vial Rack areas in the order that you want the samples to be injected.
Clicking a position turns it green. The first position that you click becomes the initial position in the sequence table. The sequence editor ignores the option selected in the
Sample Selection Layout area and fills in the remaining rows in the order of your selections.
3. Click OK to return to the New Sequence Template dialog box.
To select contiguous vials or wells in sequential order by using the Select button
1. In the Sample Positions (Click on Plate or Select Button) area, click Clear All.
This clears the Initial Position box and any selected positions on the Plate Rack and the
Vial Rack.
2. In the Sample Selection Layout area, select either the Row First or the Column First option.
3. In the Number of Samples to Select box, type the number of sample vials or wells.
4. Click Select.
The Plate Rack area displays the selected positions in green.
5. Click OK to return to the New Sequence Template dialog box.
EASY-nLC 1200 User Guide for the Xcalibur Data System
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5
Creating and Running Sequences
Running Sequences
Running Sequences
After you create a sequence with the appropriate data file names, instrument methods, and injection positions, you can run one or more rows of the sequence from the Sequence Setup view.
To run a single sample, a range of samples, or the entire sequence set
1. In the Sequence Setup view, do one of the following:
• To inject a single sample, select the specific row in the sequence table. Then choose
Actions > Run This Sample, or click the Run Sample icon, , in the Sequence
Editor toolbar.
• To inject a range of samples, select the samples that you want to inject in the sequence table. Then choose Actions > Run Sequence, or click the Run Sequence icon, , in the Sequence Editor toolbar.
–or–
• To inject the entire sequence, choose Actions > Run Sequence, or click the Run
Sequence icon, , in the Sequence Editor toolbar.
One of the following actions occurs:
• If the data system detects a new instrument configuration, the Change Instruments
In Use dialog box opens ( Figure 19 ). Go to step 2 .
• If the instrument configuration has not changed since the last sequence run, the Run
Sequence dialog box opens (
.
Figure 19. Change Instruments In Use dialog box
2. If the Change Instruments In Use dialog box opens, do one of the following:
• If the EASY-nLC instrument is not listed as the Start Instrument, click the
EASY-nLC row in the Start Instrument column and select Yes. Then click OK to accept the new setting.
• If the EASY-nLC instrument is listed as the Start Instrument, click OK.
The Run Sequence dialog box opens (see Figure 20 ).
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5
Creating and Running Sequences
Running Sequences
Start instrument
Figure 20. Run Sequence dialog box
Thermo Scientific
3. Check whether the EASY-nLC instrument is listed as the start instrument in the
Acquisition Options area.
The Start Instrument column displays Yes for the instrument that triggers data acquisition. When the EASY-nLC instrument begins its gradient program, it signals the
MS detector to begin data acquisition. This signal is transmitted through the contact closure cable.
4. Depending on whether the EASY-nLC instrument is listed as the start instrument, do one of the following:
• If it is listed as the start instrument, go to
.
• If it is not listed as the start instrument, go to
.
5. To make the EASY-nLC instrument the start instrument, do the following: a. Click Change Instruments.
The Change Instruments In Use dialog box opens.
b. Click the Thermo EASY-nLC row in the Start Instrument column.
The blank cell now displays Yes.
c.
Click OK to accept the new setting and close the dialog box.
6. In the Run Sequence dialog box, do the following: a. Make sure that the Start When Ready check box is selected.
b. Check the range in the Run Rows box.
c.
Click OK to submit the sample run to the acquisition queue.
If the Automatic Pre-run Maintenance feature is turned on and more than 8 hours have elapsed since the last sequence began, the instrument automatically runs the Flush Air script before running the new sequence. For more information about the Automatic Pre-run
Maintenance feature, see “Modifying the HPLC Properties” on page 20
.
EASY-nLC 1200 User Guide for the Xcalibur Data System
47
5
Creating and Running Sequences
Taking Corrective Action for Run-Time Errors
Taking Corrective Action for Run-Time Errors
The method editor in the EASY-nLC view of the Instrument Setup window checks each parameter entry; however, it does not check for volume conflicts among the entries.
If you have specified an invalid volume for sample loading, column equilibration, or the gradient program, an error message appears when you attempt to run samples with the instrument method from the Sequence Setup view.
describes the corrective action for common run-time errors that are caused by incompatible settings in the instrument method (.METH).
Table 11. Run-time errors caused by incompatible settings in the instrument method
Message
Sample pickup flow must be greater than 0 when pickup volume is not 0.
Solvent A consumption in the method is greater than 120.0 during precolumn, analytical column, and sample loading.
Corrective action
Enter a nonzero flow rate in the Flow box in the
Sample Pickup area on the Sample Pickup and
Loading Page.
Modify the method so that the total volume is less than 120 μL for these three steps: sample loading, precolumn equilibration, and analytical column equilibration.
Gradient: Solvent A consumption is greater than 132 μL.
Autosampler wash flush volume out of range:137 (30.25<—>100.75)
Check the gradient table. Reduce the volume of solvent A in the gradient program.
For a 20 μL sample loop, enter a volume from 30.25 to 100.75 μL in the Flush Volume box in the
Standard Wash area of the Autosampler page.
lists a common run-time error caused by incorrect entries in the Xcalibur sequence.
Table 12. Run-time error caused by incorrect entries in the sequence table
Message
EASY-nLC devices reported an error during the Prepare for Run command.
Remedy
Reduce the injection volume (sample pickup volume).
The sample pickup volume cannot be greater than the sample loop size minus 2 μL, or 18 μL for a 20 μL sample loop.
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6
Viewing the Run Status in the Information View
Use the Status page of the Xcalibur Information view to view the run status of the
EASY-nLC instrument.
To monitor the run status for the EASY-nLC instrument from the Information view
1. Open the Xcalibur Home Page window as follows:
• From the Processing Setup window, choose GoTo > Xcalibur Home Page.
• From the Instrument Setup window, click the Return to Home Page icon, , on the toolbar.
Note The Qual and Quan Browser windows open in front of the Home Page window.
2. If the Information view is hidden, click the Info View icon, , in the View toolbar
).
3. During a run, click the Thermo EASY-nLC expand/collapse icon to display the
EASY-nLC run progress and the instrument component readouts on the bottom portion of the Status page.
4. If the MS detector remains in the Waiting for Contact Closure state during a run, do the following:
• Check the contact closure connection between the contact closure pins on the back panel of the EASY-nLC instrument and the MS detector’s Start In pins.
• Check the MS detector selection (with the part number for the appropriate contact closure cable) on the Configuration > Connections page of the EASY-nLC touch-screen application.
Figure 21 shows the Status page for an LC/MS system with an EASY-nLC instrument and an
LTQ XL™ MS detector. The Load Sample state for the EASY-nLC (instrument) means that the EASY-nLC instrument is currently loading the sample onto the column assembly. The
Waiting for Contact Closure state for the mass spectrometer means that the MS detector is waiting for the contact closure signal from the EASY-nLC instrument.
Thermo Scientific EASY-nLC 1200 User Guide for the Xcalibur Data System
49
6
Viewing the Run Status in the Information View
Figure 21. Status page of the Information view
Info view icon
Expand/collapse icon
Status messages
Run-time steps for the EASY-nLC instrument
During a run, the MS detector remains in the Waiting for Contact Closure state until the LC instrument sends the start data acquisition signal.
Status of the instrument components
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EASY-nLC 1200 User Guide for the Xcalibur Data System Thermo Scientific
I
Index
A
About box, touch-screen application
Acquisition Server error messages
Automatic Pre-run Maintenance feature
autosampler model
Autosampler page
B
blank injections
C
Change > Tray Name command
Change Instruments In Use dialog box
Column first option, Sample Position Selector dialog box
column heater, EASY-Spray ion source
communication issues caused by Ethernet cable or incorrect IP addresses
caused by virus scanner software
testing the Ethernet connection
computer
configuration parameters, EASY-nLC
contact closure cable
contacting us
current user, touch-screen application
Custom option, autosampler wash step
Cycles box, autosampler wash step
D
data acquisition, premature termination of
data system computer connecting to EASY-nLC instrument
power saver settings
system requirements
virus scanner setting
device driver
Thermo Scientific installing
system requirements
uninstalling
Device Types list
Devices InstallShield Wizard
Driver Info readout
duration, gradient program
E
EASY-nLC VI control software
Autosampler page
EASY-nLC Configuration dialog box
EASY-nLC view, Instrument Setup window
Gradient page
Pre-Column and Analytical Column page
Sample Pickup and Loading page
Sample Position Selector dialog box
,
EASY-Spray column heater adding to the Devices list
controlling the temperature
Edit button, EASY-nLC Configuration dialog box
error messages, run time
Ethernet connection
F
F1 key, accessing Help with
firewall, network
flow rate analytical column equilibration step
gradient program
precolumn equilibration step
sample loading step
sample pickup step
Foundation platform
G
Gradient page
EASY-nLC 1200 User Guide for the Xcalibur Data System 51
Index: H
H
Heater port, EASY-Spray ion source
Help, accessing
hibernate setting
Home Page icon
I
Idle Flow Rate readout
Idle Mixture readout
Information view, Xcalibur data system
Initial Position entry
injection volume
instrument control software
See device driver and EASY-nLC VI control software
Intelligent Flow Control (IFC) system readout
IP address, EASY-nLC instrument
IRC communication
L
LAN port, EASY-nLC
Loop Volume readout
M
manuals, accessing from the data system computer
maximum pressure analytical column equilibration step
precolumn equilibration step
sample loading step
McAfee virus scanner
microwell plates
MS detector, triggering data acquisition
N
negative value in the Inj Vol column of the
Sequence Setup view
network cards
network configuration, EASY-nLC
network firewall
New Sequence Template dialog box, Xcalibur
nomenclature, sample position
P
parameter description tables
Autosampler page
EASY-nLC Configuration dialog box
EASY-Spray page
Gradient page
Pre-column and Analytical Column page
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EASY-nLC 1200 User Guide for the Xcalibur Data System
Sample Pickup and Loading page
Sample Position Selector dialog box
P-Bus cable
plate installed, selecting
Plate Rack area, Sample Position Selector dialog box
plate type
port 22, TCP/IP traffic
Position column, Sequence Setup view table
power saver settings
Pre-Column and Analytical Column page
prerequisites, installation
pressure, maximum
pump solvents, A and B
R
readouts
EASY-nLC Configuration dialog box
Status page, Information view
Tray Type box, New Sequence Template
dialog box
release notes
requirements computer
software
Row First option, Sample Position Selector dialog box
Run Sample icon, Sequence Setup view
Run Sequence dialog box
Run Sequence icon, Sequence Setup view
run time error messages
S
sample layout and start position, configuration setting for
sample layout, selecting
sample loop size
Sample Pickup and Loading page
Sample Position Selector dialog box selecting sample layout and start position for the instrument configuration
selecting the vial or microplate well positions for a sequence
screen saver settings
Select Vials button, New Sequence Template dialog box
sequence runs premature termination of
setting up the injection sequence table
starting from the Sequence Setup view
Sequence Setup view of the Home Page window
software requirements
solvents
Thermo Scientific
specifying in solvent reservoir bottles
wash bottles W1 to W3
Source list, autosampler wash step
Standard option, autosampler wash
Start Position readout
Status page of the Information view, Home Page window
system requirements
T
temperature, EASY-Spray source column heater
Test Connection button
Thermo EASY-nLC Configuration dialog box
Thermo Foundation Instrument Configuration window
Thermo logo, touch-screen application
touch-screen application
About box
Maintenance > Devices page
selecting the MS detector connected through contact closure to the EASY-nLC instrument
version
tray type
troubleshooting communication issues
,
contact closure issues
–
data acquisition issues
run-time errors
U
URLs, consumable parts
User Name configuration parameter
user name, default
V
Vial Rack area, Sample Position Selector dialog box
vial rack positions
vial tray
virus scanner, McAfee
volume analytical column equilibration
column in custom wash table
conflicts
flush, autosampler wash step
precolumn equilibration
sample loading
sample pickup
W
waiting for contact closure
wash, autosampler
Thermo Scientific
Index: T
Windows updates, automatic
X
Xcalibur data system
Column Arrangement dialog box, Sequence Setup
view
Instrument Setup window
New Sequence Template dialog box
Sequence Setup view
Status page, Information view, Home Page window
EASY-nLC 1200 User Guide for the Xcalibur Data System
53
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Table of contents
- 3 Contents
- 5 Preface
- 6 Related Documentation
- 7 System Requirements
- 8 Data System Computer Settings
- 9 Cautions and Special Notices
- 10 Contacting Us
- 11 Installing the EASY-nLC Control Software
- 11 Checking the System Prerequisites
- 12 Removing a Previous Version of the Instrument Control Software
- 13 Installing the Current Version of the Instrument Control Software
- 14 Excluding the EASY-nLC from the VirusScan Enterprise
- 17 Setting Up the Foundation Instrument Configuration
- 17 Connecting the EASY-nLC Instrument to the Data System Computer
- 18 Adding the System Devices to the Instrument Configuration
- 19 Setting Up the EASY-nLC Configuration
- 25 Setting Up the Sample Layout and Start Position
- 27 Modifying the HPLC Properties and Controlling Other Devices
- 27 Accessing the Devices Page of the Touch-screen Application
- 28 Adding the EASY-Spray Device to the Devices List
- 30 Modifying the HPLC Properties
- 33 Creating Xcalibur Instrument Methods
- 33 Setting Up the EASY-nLC Instrument Method Parameters
- 35 Sample Pickup and Loading Page
- 37 Gradient Page
- 40 Pre-Column and Analytical Column Page
- 43 Autosampler Page
- 46 EASY-Spray Page
- 48 Saving Instrument Methods
- 49 Creating and Running Sequences
- 49 Creating Sequences
- 53 Setting Up the Injection Positions with the Sample Position Selector
- 56 Running Sequences
- 58 Taking Corrective Action for Run-Time Errors
- 59 Viewing the Run Status in the Information View
- 61 Index