Pathfinder 1.0 Administrator Guide Version A

Pathfinder 1.0 Administrator Guide Version A
Pathfinder
Version 1.0
Administrator Guide
XCALI-97242 Revision A
June 2009
© 2009 Thermo Fisher Scientific Inc. All rights reserved.
Xcalibur is a registered trademark of Thermo Fisher Scientific Inc. in the United States.
Windows, Excel, and Windows Vista are registered trademarks of Microsoft Corporation in the United States
and other countries. Adobe, Acrobat, and Reader are registered trademarks of Adobe Systems Incorporated in
the United States and other countries.
Pathfinder is a trademark of Thermo Fisher Scientific Inc.
All other trademarks are the property of Thermo Fisher Scientific Inc. 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.
Thermo Fisher Scientific Inc. makes no representations that this document is complete, accurate or errorfree and assumes no responsibility and will not be liable for any errors, omissions, damage or loss that might
result from any use of this document, even if the information in the document is followed properly.
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: Pathfinder Version 1.0.
For Research Use Only. Not for use in diagnostic procedures.
C
Contents
Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .v
Related Documentation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .vi
System Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .vi
Contacting Us . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vii
Thermo Scientific
Chapter 1
Getting to Know Pathfinder . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1
Understanding Pathfinder . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
How Pathfinder Works . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
Quick Access to Pathfinder Tasks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4
Using the Pathfinder Administrator Application . . . . . . . . . . . . . . . . . . . . . . . . . 5
Chapter 2
Getting Started with Pathfinder . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .9
Opening Pathfinder. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Analyzing a Sample . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Analyzing Groups of Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Viewing Reports . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
Chapter 3
Working with Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .17
Working with Instrument Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Creating an Instrument Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Changing an Instrument Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Working with Pathfinder Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
Creating a Pathfinder Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
Changing a Pathfinder Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
Working with Method Groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44
Creating a Method Group . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44
Modifying a Method Group . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46
Changing the Active Method Group . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
Deleting a Method Group . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
Chapter 4
Working with Samples. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .49
Submitting a Single Sample for Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
Submitting Multiple Samples for Analysis Manually . . . . . . . . . . . . . . . . . . . . . 52
Creating Files for Importing Lists of Multiple Samples . . . . . . . . . . . . . . . . . . . 54
Submitting an Imported List of Samples for Analysis . . . . . . . . . . . . . . . . . . . . 55
Viewing Instrument Status During Processing. . . . . . . . . . . . . . . . . . . . . . . . . . 56
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Contents
Chapter 5
Viewing Workflow Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .59
Viewing Data Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
Viewing Results for a Workflow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
Viewing Peak Details. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64
Viewing Specific Details . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65
Viewing Reports . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
Modifying Sample Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74
Working with peaks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74
Reprocessing Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76
Changing Parameter Values for a Single Sample . . . . . . . . . . . . . . . . . . . . . . 76
Reprocessing a Group of Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77
Chapter 6
Managing Users and the Pathfinder System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .79
Working with Users . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80
Adding Users . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80
Changing User Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
Removing a User . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 81
Defining System Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82
Configuring Pathfinder on Remote Workstations . . . . . . . . . . . . . . . . . . . . . 83
Setting Up a Virtual E-Mail Server . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85
Configuring Instruments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
Initializing Pathfinder Using Xcalibur . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 92
Setting Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 93
Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98
Appendix A Pathfinder Reference . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .99
Pathfinder Menus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
File Menu . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
Tools Menu. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100
Go Menu. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101
Help Menu . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101
Method Development View . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 102
Method View . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 102
Instrument Configuration Views . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 116
Method Group Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Analysis View . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Data Review Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118
Reports Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
Real Time Status View . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
User Administration View . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
User View Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
User Mode Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
Survey Screening Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
Target Matching Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 123
Target Screening Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 124
Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .125
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Preface
This guide describes how to use Thermo™ Pathfinder™ to analyze a variety of samples.
Contents
• Related Documentation
• System Requirements
• Contacting Us
Y To suggest changes to documentation or to Help
Complete a brief survey about this document by clicking the link below.
Thank you in advance for your help.
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Preface
Related Documentation
Pathfinder includes Help and these manuals as PDF files:
• Pathfinder Quick Start
• Pathfinder Administrator Guide
Y To view product manuals
Go to Start > Programs > Thermo Pathfinder 1.0 > Manuals.
Y To open Help
• From the Pathfinder window, choose Help > Contents.
• In a specific window or dialog box, click Help or press F1 for information about
setting parameters.
For more information, including upcoming application notes, visit
www.thermo.com/appnotes.
System Requirements
In addition, your system that supports online instruments must meet these minimum
requirements.
System
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Pathfinder User Guide
Requirements
PC
•
•
•
•
•
2 GHz processor with 2GB RAM
CD-ROM drive
Video card and monitor capable of 1280× 1024 resolution (XGA)
75 GB or greater available on the C: drive
NTFS format
Software
• Microsoft™ Windows™ XP Professional with Service Pack 3
• Microsoft Office 2003 or 2007
• Xcalibur™ 2.1
Thermo Scientific
Preface
Contacting Us
There are several ways to contact Thermo Fisher Scientific for the information you need.
Y To contact Technical Support
Phone
800-532-4752
Fax
561-688-8736
E-mail
us.techsupport.analyze@thermofisher.com
Knowledge base
www.thermokb.com
Find software updates and utilities to download at mssupport.thermo.com.
Y To contact Customer Service for ordering information
Phone
800-532-4752
Fax
561-688-8731
E-mail
us.customer-support.analyze@thermofisher.com
Web site
www.thermo.com/ms
Y To copy manuals from the Internet
Go to mssupport.thermo.com and click Customer Manuals in the left margin of the
window.
Y To suggest changes to documentation or to Help
• Fill out a reader survey online at www.thermo.com/lcms-techpubs.
• Send an e-mail message to the Technical Publications Editor at
techpubs-lcms@thermofisher.com.
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1
Getting to Know Pathfinder
Pathfinder is a data acquisition, processing, and reporting system for users who have a range of
experience levels with liquid chromatography mass spectrometry (LC-MS).
This chapter provides a basic overview of Pathfinder and demonstrates how you can use
Pathfinder in the work environment. For information about configuring Pathfinder, see
“Defining System Information” on page 82.
Contents
• Understanding Pathfinder
• How Pathfinder Works
• Quick Access to Pathfinder Tasks
• Using the Pathfinder Administrator Application
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Getting to Know Pathfinder
Understanding Pathfinder
Understanding Pathfinder
You can access Pathfinder in one of two different environments, as Administrator or User.
These names represent two different and nonoverlapping access modes. The following table
describes what each account name allows.
Table 1. End-user Modes for Pathfinder
Account name
Tasks and permissions
Administrator
• Develops and tests methods for sample analysis with
manual and automated data processing.
• Develops data review and reporting formats.
• Analyzes sample run data.
• Manages the user mode accounts.
User
• Submits samples for analysis.
• Receives the results of automated sample analysis and
data processing.
You can use Pathfinder to develop and save multiple sets of experimental definitions
(methods) and related information. Then you can run these saved instruction sets to
automatically process and report on LC-MS data. Pathfinder works with high-mass accuracy
and high-resolution data sets, such as elemental composition for data analysis, reduction,
review, and reporting.
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Getting to Know Pathfinder
How Pathfinder Works
How Pathfinder Works
Figure 1 illustrates the workflow to use when setting up and analyzing your data. It includes
the workflow for both administrators and users. Click workflow entries in bold to go to the
section describing that process.
Figure 1.
Pathfinder workflow overview for both Administrator and User accounts
LC-MS technique produces raw data
Pathfinder
Administrator mode
Specify general method
information
Define peak processing
User mode
Click workflow
Select method
Define MS processing
Enter user and sample ID
Define report settings
Test method
Enter sample file name
(Enter target molecular
weight)
Save and distribute
method
Click Go to run sample
Analyze results
Collect printout
View spectra graph and
reports
Thermo Scientific
Note Optional steps are italicized.
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1
Getting to Know Pathfinder
Quick Access to Pathfinder Tasks
Quick Access to Pathfinder Tasks
The following table shows you the location in Pathfinder to perform tasks in Administrative
mode:
Task
Task location in Pathfinder
To run a sample
See “Getting Started with Pathfinder” on page 9.
To create or change a processing method
See “Working with Pathfinder Methods” on
page 21.
To specify general method information
Click Method Development. Select New
method or Open Method in the Method
Development view. Click the General tab.
To define processing values for a Target
Screening method
Select Target Screening as a workflow on the
General page and click the MS Processing tab.
To define peak processing for Target
Select Target Matching or Survey Screening as a
Matching and Survey Screening methods workflow on the general page and click the Peak
Processing tab.
To define MS processing for Target
Click the MS Processing tab.
Matching and Survey Screening methods
To define report settings for Target
Click the Reports tab.
Matching and Survey Screening methods
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To save method
Click Save method in the Method Development
view.
To test method
Click User Administration in the Method
Development view. Select a test type under User
Mode in the User Administration view.
To view results
See “Viewing Workflow Results” on page 59.
To view test results
Click Analysis in the Method Development
view. Click Open sequence. Click Data review
or Reports view to verify test data.
To configure instruments
See “Configuring Instruments” on page 89.
To add, open, or save instrument
methods to configure instruments
Click Method Development. Click Instrument
Methods in the Method Development view.
To add or change method groups
“Working with Method Groups” on page 44.
To add, modify, activate, or delete
method groups
Click Method Development. Click Method
Groups in the Method Development view.
Thermo Scientific
1
Getting to Know Pathfinder
Quick Access to Pathfinder Tasks
Thermo Scientific
Task
Task location in Pathfinder
To add or change users
See “Working with Users” on page 80.
To add, modify, delete users
Click User Administration in the Method
Development view. Click User View in the User
Administration view.
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1
Getting to Know Pathfinder
Using the Pathfinder Administrator Application
Using the Pathfinder Administrator Application
This section describes the main sections of the Pathfinder Administration GUI.
After you log into Pathfinder, the main window opens.
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1 Getting to Know Pathfinder
Using the Pathfinder Administrator Application
This page displays three main buttons.
Icon
Description
Areas
Opens the Method Development
view where you can do the
following:
• Create, modify, and save
instrument methods
• Create, modify, and save
processing methods.
• Define and activate method
groups
Click the button at the left of a view
to hide the options. Click the name
of a view to display the options.
Opens the Analysis view where you
can do the following:
• View real time status and
instrument processing
• Open, save, or reprocess a
sequence
• View data information for a
specific sequence
• View reports for a sequence
Click the button at the left of a view
to hide the options. Click the name
of a view to display the options.
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Icon
Description
Opens the User Administration
view where you can do the
following:
• Process sequences using the
User Mode
• Create, modify, and delete users
Click the button at the left of a view
to hide the options. Click the name
of a view to display the options.
Areas
2
Getting Started with Pathfinder
Pathfinder guides you in the analysis of one or more LC-MS samples using predefined
methods to produce automated reports. This quick start chapter describes how to submit
samples in User mode.
Contents
• Opening Pathfinder
• Analyzing a Sample
• Analyzing Groups of Samples
• Viewing Reports
Click the icon below to see a video explaining how to process data.
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2
Getting Started with Pathfinder
Opening Pathfinder
Opening Pathfinder
Y To open Pathfinder
1. Choose Start > Programs > Thermo Pathfinder 1.0 > Pathfinder or double-click the
Pathfinder icon on your computer desktop. The Pathfinder logon screen opens.
2. Click User. The Pathfinder User window opens.
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Getting Started with Pathfinder
Analyzing a Sample
Analyzing a Sample
Use one of the Pathfinder workflows to submit a sample for analysis. You must define a
processing method before the user can analyze a sample (see “Creating a Pathfinder Method”
on page 21).
Y To run a single sample
1. From the Pathfinder User window, choose a workflow.
• To view peak results, click Survey Screening. Adjust peak processing values to define
an area of interest.
• To search for a target compound with a specified molecular weight, click Target
Matching.
• To search for several compounds in a library, click Target Screening.
The page for that workflow opens.
2. Type a unique personal identifier in the User name box. Pathfinder uses this ID to notify
you when reports are available.
3. Type the sample identification in the Sample ID box.
4. Type the autosampler vial position in the Vial position box.
If the vial position value is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Go. For more information about the
autosampler vial position format, see your autosampler documentation.
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2
Getting Started with Pathfinder
Analyzing a Sample
5. Type the name of the sample data file in the File name box, unless Pathfinder grays out
the file name.
6. If the workflow is Target Matching, type the monoisotopic molecular weight that you
want to match in the Target MW box.
The molecular weight entered must be the monoisotopic molecular weight and not the
average molecular weight.
7. Select a method from the Method area. When you click a method name, Pathfinder
displays a method description if available to help you choose the appropriate method.
8. Click Go to start the sample analysis. If the method specifies a printed report, Pathfinder
prints a report using the printer selected in the method.
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Getting Started with Pathfinder
Analyzing Groups of Samples
Analyzing Groups of Samples
Y To run a group of samples using an imported sample list
1. From the Pathfinder User window, choose a workflow. The page for that workflow opens.
2. Type a user name in the User name box.
3. Select a specific method from the Method area.
4. Click Import to add a file containing sample information to the method. A browse
window opens. Browse to where your sample files are stored. Sample files are initially
stored in C:\Thermo\Pathfinder\Templates.
5. Select the samples file and click Open. In the Sample ID table, make sure each sample has
the correct vial position.
6. To change the information on any sample, select the sample and enter data in the boxes to
the left. Click Update to save the changes.
7. Click Go. If the method specifies a printed report, Pathfinder prints it using the printer
selected in the method.
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2
Getting Started with Pathfinder
Analyzing Groups of Samples
Y To analyze a group of samples using a manually created sample list
1. From the Pathfinder User window, choose a workflow. The page for that workflow opens.
2. Type the user name in the User Name box.
3. Select a specific method from the Method area.
4. Enter sample information:
a. Type the first sample ID in the Sample ID box.
b. Type the first autosampler vial position in the Vial position box.
c. Type the file name of the sample data file in the File name box, unless Pathfinder
supplies the name. Each sample must have a unique file name.
d. If you chose Target Matching, type a molecular weight in the Target MW box. The
molecular weight entered must be the monoisotopic molecular weight. It cannot be
the average molecular weight.
e. Click Add. The sample information is displayed in the table to the right.
5. Type information for the remaining samples, repeating all of step 4 for each sample.
6. To change the information on any sample, select the sample and enter data in the boxes to
the left. Click Update to save the changes.
7. Click Go. If the method specifies a printed report, Pathfinder prints it using the printer
selected in the method.
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2
Getting Started with Pathfinder
Viewing Reports
Viewing Reports
When viewing a Target Screening report, look for the following:
• In the summary report, a table of all compounds from the library with the retention times
when they were found, the m/z value found, the difference from the expected value, and
an intensity calculation. Also, information on the adducts found and any matching HCD
fragments.
• In the long report, all the information from the summary report for each compound.
Also, if adducts are identified, the mass chromatogram for each adduct and, if matching
fragments are found, the mass chromatogram for the fragment.
There is an option to generate reports on only found compounds or on all compounds in the
user library.
When viewing a Survey Screening report, look for the following:
• Information on each peak found including a chromatogram that highlights the peak
found, a mass spectrum of that chromatographic peak, HCD spectrum (if applicable),
and area and intensity calculations for that peak.
• A list of calculated formulas for the most intense peak in the mass spectrum.
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2
Getting Started with Pathfinder
Viewing Reports
When viewing a Target Matching report, look for the following:
• Information on any peaks that match the target m/z.
• For each match, a chromatogram showing the match, the mass spectrum, the HCD
spectrum (if applicable), and area and intensity calculations for the matched peak. Also, a
graphical representation of the peak area integration.
• A list of calculated formulas for the target match candidate from the mass spectrum.
• A table on the first page of the target matching report that shows all peaks found for the
target mass. If the peak matches the boundary conditions specified in the method, then
Pathfinder displays Yes in the match column.
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3
Working with Methods
A Pathfinder method represents the set of instructions for the acquisition, processing, and
reporting to be performed by the LC/MS system on a sample submitted for analysis. This
chapter describes how to create and modify methods.
Contents
• Working with Instrument Methods
• Working with Pathfinder Methods
• Working with Method Groups
Working with Instrument Methods
An instrument method defines the data acquisition parameters for a sample, resulting in a
data file that can then be processed using the other elements of a Pathfinder method. You can
use this data file in other software applications, such as Xcalibur’s Qual Browser.
Use the Instrument Method view to change the options and parameters for each of the devices
that are available as part of the system configuration. The devices include the
autosampler (AS), liquid chromatograph (LC), and mass spectrometer (MS).
You can create new instrument methods, modify existing instrument methods, and save
instrument method files. When you define an instrument method, it resides on the
instrument where you define the method.
Note Before you use the Instrument Method View, use the Instrument Configuration
program to select the instrument and devices for your analyses. For more information, see
“Configuring Instruments” on page 89.
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Working with Instrument Methods
On the left, the Instrument Method View displays the icons of all available devices you have
selected using the Instrument Configuration window. Each tab in the horizontal tab bar opens
the setup parameters for a specific available device.
To enter the setup parameters for a particular instrument, click the tab for that device. The
Instrument Method View then displays one or more pages of parameters that you can set.
Use the following steps to create or change an instrument method. You must create at least
one instrument method before running samples.
• Creating an Instrument Method
• Changing an Instrument Method
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Working with Instrument Methods
Creating an Instrument Method
Use an instrument method to define the data acquisition options and parameters that are
available on the individual devices that make up the analytical system, that is, the
autosampler, HPLC pump, mass spectrometer and so forth.
Y To create an instrument method
1. In the Method Development area, click New instrument method in the Instrument
Method view.
2. Click an instrument device tab on the horizontal tab bar. Pathfinder opens the method
editor for that device in the workspace and displays options related to the specific
requirements and capabilities of the device. For help setting parameter values, see the
documentation FOR that hardware device.
If the device for the instrument method you want to define is not on the list, make sure
the device has been added to the system. See “Configuring Instruments” on page 89.
3. To set parameter values, repeat steps 2 and 3 for each device on your configured system.
When finished, click Save instrument method in the Method Development area.
4. Type a name in the Save dialog box and click OK.
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Working with Instrument Methods
Changing an Instrument Method
You can change an instrument method after creating it to optimize the data acquisition
conditions.
Y To change an instrument method
1. In the Method Development mode, click Open instrument method in the Instrument
Method view. The Open Method dialog box opens.
2. Select an existing method. The Instrument Method view opens, displaying the list of
configured devices in a horizontal tab and the method editor in the workspace. Select an
instrument tab to open the method page for a specific instrument.
3. Change parameter values for any of the devices and click Save instrument method in the
Method Development area.
4. To save this modified method under a new name, choose File > Save as.
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Working with Pathfinder Methods
Working with Pathfinder Methods
A Pathfinder method, created in the Method View, specifies the instrument method used for
data acquisition and defines the data processing and reporting parameters for a sample. When
you process a sample, Pathfinder provides a set of result files and, if you specify it, a set of
electronic and hardcopy reports. Review these result files using Pathfinder's Analysis mode.
The Method View displays the options and parameters to be used for each of the data
processing steps that are available as part of the overall workflow configuration. These include
descriptive information, data acquisition, peak processing, mass spectrometer-specific
processing, and reporting. You can create new Pathfinder methods, modify existing methods,
and save method files.
Follow these major steps to create or change a Pathfinder method. You must create at least one
Pathfinder method before running samples.
Contents
• Creating a Pathfinder Method
• Creating a Processing File for Target Screening
• Changing a Pathfinder Method
• Working with Method Groups
Creating a Pathfinder Method
To create a Pathfinder method, complete these procedures:
• Defining General Values for a Method
• Defining Processing Values for Target Matching and Survey Screening
• Defining Target Screening Settings
• Choosing Report Types
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Defining General Values for a Method
Pathfinder requires some information, such as the selection of an instrument method, when
using a Pathfinder method. Other information, such as the assay type, is optional. Use the
General tab of the Method View to enter these values.
Y To enter general information about the method
1. Double-click the Pathfinder icon on your desktop or choose Start > All Programs >
Thermo Pathfinder 1.0 > Pathfinder 1.0.
2. Click Administrator on the opening screen and enter the password.
• For access to the Administrator mode, type your system password (thermo is the
default password and might be different on your system. Contact your system
administrator if the password is different.).
3. Select Method Development on the left side of the window.
4. Select New method. Method View opens to the General Page.
5. Type the name of the lab in the Lab name box.
6. Type the name of the test in the Assay type box.
7. To define the amount of the sample used for analysis, click the up and down arrows in the
Injection Volume (μL) box or type in a specific value. Enter a value from .1 to 100.
8. Select an instrument method from the Instrument method box. For more information
about instrument methods, see “Creating a Pathfinder Method” on page 21.
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9. To select a processing workflow, select from the Workflow list:
• To perform overall peak processing and elemental composition (for the mass
spectrometer data), click Survey Screening.
• To perform overall peak processing and elemental composition (for the mass
spectrometer data), and then compare each MS detected peak to a user-supplied
target molecular weight with a goal of finding a target compound in a sample, click
Target Matching.
• To find compounds in a database in the sample, click Target Screening.
To specify descriptive information about this method, enter text into the Description box.
Users will see this information when selecting a method for submission of a sample or a set of
samples.
Defining Processing Values for Target Matching and Survey Screening
Use the following steps to define processing values for Target Matching and Survey Screening.
For a Target Screening workflow, see “Creating a Processing File for Target Screening” on
page 41.
• Setting Peak Processing Values
• Defining Peak Detecting and Integration Values
• Setting High (ICIS) Integration Values
• Setting Standard (Genesis) Integration Values
• Specifying MS Processing Values
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Setting Peak Processing Values
On the Processing page in the Peak Processing view, you can select one or more data sources to
be used for peak detection. Use the other pages to define specific peak detection parameters
for the data sources selected. If you choose the Target Matching workflow, Pathfinder hides
peak processing options because they are selected using a different view. Options that are
grayed out are not available for your selected workflow.
Y To detect peaks
1. Click the Peak Processing tab to define peak parameters. The Processing Page opens.
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2. In the Data Source area, select the data traces Pathfinder will process for peak detection.
For each of these detector types, you might have one or multiple channels of data that
need to be processed, stored, and visualized.
• Select an MS option to designate one or more data traces for Pathfinder to process for
each of the available scan filters from the MS detector. When processing negative and
positive data, you can get elemental composition results for the positive mode or the
negative mode, but not both. To get elemental composition results for both the
negative and positive mode, process the modes separately, being sure to set the correct
charge state in elemental composition for each polarity. Processing an MS data source
results in a peak list with associated mass spectra:
–
To process a specific mass range, select the Mass Range check box. Use this
option with a Target Matching workflow only.
–
To process a total ion current [TIC] chromatogram, select the TIC check box.
Use this option for a Survey Screening workflow.
–
To process a base peak chromatogram, select the Base Peak check box.
• For PDA data, valid trace types are Total Scan or Spectrum Maximum. Processing a
PDA data source results in a peak list with associated PDA spectra:
–
To process the data trace for the summed wavelength intensities, select the Total
Scan check box.
–
To process the data trace for the spectrum maximum, select the Spectrum
Maximum check box.
• To select UV detector data, select a channel in the UV channel area. For UV detector
data, Pathfinder supports four channels (labeled Channel A-D). UV results only
provide a peak list.
• To select data, select from the A/D Card area. For data from an A/D card, Pathfinder
supports two channels (labeled A/D 1-2). A/D results only provide a peak list.
3. To set delay offsets:
• To set the offset for PDA data, type a value in minutes in the PDA offset (min) box.
• To set the offset for UV data, type a value in minutes in the UV offset (min) box.
• To set the offset for analog data, type a value in minutes in the Analog offset (min)
box.
• To set the offset for A/D data, type a value in minutes in the A/D offset (min) box.
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If you selected a Target Matching workflow, Pathfinder also displays options for target
matching.
4. For the targeting range, select an adduct in either the Positive ion adducts or Negative ion
adducts area to generate a target m/z value. You can select more than one adduct. The
most common adduct is H.
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Pathfinder calculates the m/z of the ionized molecular ion with the specified adduct. This
step is necessary to compensate for the difference between the measured m/z value of an
ion for a particular compound (resulting from the ionization process) and that
compound’s non-ionized molecular weight.
For example, in LC-MS using positive polarity API, the ion can be formed by the
addition of a positive charge (proton) to the compound. In this instance, the observed
ion’s m/z value for that compound is shifted by 1.0073 amu relative to its molecular
weight. Since the values that you submit for target matching are given as the
monoisotopic molecular weight of the compound of interest, the measured ion m/z value
must have the value of any adduct present subtracted from it to determine the observed
non-ionized molecular weight and make the appropriate comparison to the target
molecular weight.
Define the value to add or subtract by selecting one or more adducts from the available
list or by specifying a custom adduct if the choice is not available (for example, a unique
LC mobile phase chemistry).
Pre-defined adducts and their adjustments include the following:
Positive ion
Negative ion
Adduct
Adjustment
Adduct
Adjustment
H
+ 1.0073
H
- 1.0073
H3O
+ 19.0178
NH4
+ 18.0338
Na
+ 22.9892
K
+ 38.9632
5. Define the width of the mass window. Pathfinder uses your value above and below the
mass. The default is 5 ppm. Pathfinder uses this value for the mass tolerance for target
matching.
6. If you are specifying details for a Target Matching workflow, set a value for the Minimum
acceptable relative intensity by entering a value from 1 to 100. Compounds that match
the mass specified in the workflow but that fall below the intensity threshold will not
appear in the report because the peak was not defined as a match. This setting is not
available for survey screening.
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7. To enter specific values for finding peaks, in the Peak Processing view, click the tab for a
type of detector (MS, PDA, UV, or A/D Card). The page for that detector type opens.
8. Specify a sensitivity level for the peak detector algorithm by selecting from the Sensitivity
list. For information about setting peak integration values, see “Defining Peak Detecting
and Integration Values” on page 29.
• To use high sensitivity levels for detection, select High (ICIS). This setting, designed
for mass spectrometers, performs a more exhaustive peak processing pass.
• To use standard sensitivity levels for detection, select Standard (Genesis). This
setting performs a comprehensive peak processing pass.
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9. To define the retention time range for peak processing, select the Limit the retention
time range check box. Specify a start (Min RT) and end time (Max RT) for the range.
10. To turn on a peak threshold for defining the lower limit for peak retention, select the
Enable peak threshold check box.
• Define the lowest value of the threshold as a percentage of the largest peak.
• To evaluate peaks by height, select the By height option, or to evaluate them by area,
select the By area option. By area is the default option.
11. To store a defined number of peaks by size, starting with the largest peak, select the Only
select top peaks check box.
• Define the number of largest peaks to store by clicking the up or down arrow in the
Only select top peaks box. You can also type a value using the keyboard.
• To evaluate peaks by height, select the By height option, or to evaluate them by area,
select the By area option.
Defining Peak Detecting and Integration Values
Follow these procedures to set detection and integration values for algorithms that control the
sensitivity of the peak processing results. If you choose the target matching workflow,
Pathfinder hides peak detection and integration options. Options that are grayed out are not
available for your selected workflow.
• Setting High (ICIS) Integration Values
• Setting Standard (Genesis) Integration Values
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Setting High (ICIS) Integration Values
1. Select High (ICIS) in the Sensitivity list. The ICIS Peak Integration Area opens.
2. In the Smoothing points box, define the number of points to use for chromatogram
smoothing. Type an odd number from 3 for minimum smoothing to 15 for maximum
smoothing. To perform no smoothing, type 1 in the box.
3. To specify the number of scans to review, looking for a local minima, type a value in the
Baseline window box. The valid range is 1 through 500. The default value is 40 scans.
4. To define the noise level multiplier used to determine the peak edge after the location of
the possible peak, type a value from 1 through 500 in the Area noise factor box.
Pathfinder excludes peaks that are less than the selected threshold.
5. To specify the noise level multiplier used to determine the potential peak signal threshold,
type a value from 1 through 1000 in the Peak noise factor box.
6. To limit the peak width of a component during peak integration of a chromatogram,
select the Constrain peak width check box. You can then set values that control when
peak integration is turned on and off by specifying a peak height threshold and a tailing
factor.
7. To change the peak height value and test the width of target peaks, enter a value in the
Peak Height box. You can enter any value from 0 to 100 percent.
8. To control how Pathfinder integrates the tail of a peak, enter a value from 0.5 through 9.0
in the Tailing factor box. This factor is the maximum ratio of the trailing edge to the
leading side of a constrained peak.
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Setting Standard (Genesis) Integration Values
Y To set Genesis peak integration values
1. Select Standard (Genesis) in the Sensitivity list. The Genesis Peak Detection Area opens.
2. Define the number of points to use for chromatogram smoothing. Type an odd number
from 3 for minimum smoothing to 15 for maximum smoothing in the Smoothing points
box. To perform no smoothing, type 1 in the box.
3. Define the signal to noise (S/N) threshold for detecting peak edges. The default value is
0.5 and the valid range is 0.0 to 999.0. Pathfinder excludes peaks that are less than the
selected threshold.
4. To help detect and separate unresolved peaks, select the Enable valley detection check
box to use the Genesis valley detection approximation method. This method drops a
vertical line from the vertex of the valley between unresolved peaks to the baseline. The
intersection of the vertical line and the baseline defines the end of the first peak and the
beginning of the second peak.
5. To set the expected peak width parameter (in seconds), type a value in the Expected
width (sec) box. This value defines the minimum width that a peak is expected to have
and is used with the valley detection option to determine unresolved as opposed to
asymmetric peaks.
6. To limit the peak width of a component during peak integration of a chromatogram,
select the Constrain peak width check box. You can then set values that control when
peak integration is turned on and off by specifying a peak height threshold and a tailing
factor.
7. To change the peak height value and test the width of target peaks, type a value in the
Peak Height box. You can type any value from 0 to 100 percent.
8. To control how Pathfinder integrates the tail of a peak, enter a value from 0.5 through 9.0
in the Tailing factor box. This factor is the maximum ratio of the trailing edge to the
leading side of a constrained peak.
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Specifying MS Processing Values
Use the MS Processing tab of the Pathfinder method to define the unique processing
requirements for the mass spectrometer data of the sample. If you are defining processing
requirements for a Target Matching or Sample Survey workflow, select the Elemental
Composition tab to specify the options for the Spectral Fit algorithm used for determining
elemental formulas based on mass spectral ions and their intensities. Use the following
sections to define processing values for mass spectrometry processing.
Use the MS Processing: Elemental Composition Page to set values that calculate one or more
possible empirical formulas for a particular chromatographic peak. Options that are grayed
out are not available for your selected workflow.
When you process a Survey Screening sample, the base peak is the largest peak in the sample
defined by height. For a Target Matching method, Pathfinder finds the peak closest to the
value entered in the Target MW field. The resulting list of peaks is sorted according to the
spectral fit value.
Pathfinder uses the isotope patterns and intensities surrounding the base peak to provide a
more rigorous match than the standard elemental composition models. In the Elemental
Composition view, you specify spectral fit parameters. The algorithm does the following:
• Identifies the base peak (by height) in a particular peak’s mass spectrum.
• Determines the monoisotopic peak, if not the base peak.
• Examines a set of ions around the monoisotopic peak, including the monoisotopic
cluster.
• Calculates all elemental composition candidates based on a single exact mass (usually the
monoisotopic mass).
• Scores the match between the measured isotope pattern and the theoretical isotope
pattern of each composition candidate with a Fit Factor. A good fit factor (100 percent is
the maximum) is reached only if both the measured and the theoretical isotope patterns
and intensities are almost identical. If the patterns are not similar, the fit factor is 0
percent.
• Sorts the elemental composition candidates in terms of decreasing fit. The candidate with
the best fit factor is considered to be the most likely match.
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Y To define elemental composition values for survey screening and target matching
1. Click the MS Processing tab. The MS Processing: Elemental Composition Page opens.
2. Define the expected charge state by entering a value from -99 to 99.
3. Select whether or not to use the Nitrogen rule in the formula calculation. Select from Do
not use, Odd electrons, or Even electrons.
4. To restrict the number of possible elemental compositions, enter a value in the Mass
tolerance box. Pathfinder returns results of the elemental composition search only if the
computed formula matches the observed mass within the specified tolerance. Pathfinder
uses this value for mass tolerance throughout the application for elemental composition
(acceptance and monoisotopic mass determination).
5. To set the values for double bonds and ring equivalents—a measure of the number of
unsaturated bonds in a compound—and limit the calculated formulas to only those that
make sense chemically in this regard, type values in the RDB equiv low box and the RDB
equiv high box.
6. Specify the total number of results by typing a value or clicking the up and down arrows
in the Max results box.
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7. To use the standard carbon isotope distribution for elemental composition, select the Use
representative elements check box (13C abundance = 1.070%). To use the carbon
isotope distribution that is common in proteins, clear this option (13C abundance =
1.086%).
8. To select elements to use in the element formula candidate, click
to display the
periodic table. Select an element and click Apply to add the element to the list. If the
element is already in the grid, you can activate it by selecting the use check box.
• To remove elements from use in the elemental formula, select the element in the grid
and click
.You can also turn off the element by clearing the use check box.
• To define the number of elements in a calculated candidate, specify a maximum or
minimum value in the grid.
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9. To set advanced parameters, including those for spectral fit results, click Show Advanced.
The Advanced area opens. To close the Advanced area, click Hide Advanced.
10. To specify the expected mass error in the spectrum data, enter a value in the Expected
mass error box. This value is used by the algorithm. For best results, set the expected mass
error to a value that causes less than 98 percent of all mass errors to be smaller than the
expected mass error. The expected mass error must be specified in ppm (parts per
million).
11. Enter a value in the Expected intensity error box so that the algorithm knows the
expected intensity error of the mass spectrometer. For best results, set the expected error
to a value that causes less than 98 percent of all intensity errors to be smaller than the
expected intensity error value.
12. Set a penalty to identify peaks that are specifically searched for but not found. If the
theoretical isotope pattern of a candidate and the measured isotope pattern are compared,
Pathfinder might not find a measured packet for a theoretical isotope peak. This is called
a missing peak.
13. To measure an observed mass error on an isotope peak upon its abundance, select the
Mass error weighted by abundance check box. This ensures that more abundant masses
have a smaller percentage of errors than less common masses.
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14. To measure an observed intensity error on an isotope peak upon its abundance, select the
Intensity error weighted by abundance check box. More abundant intensity readings
have a smaller percentage of error than less common ones.
15. To measure intensity errors by absolute value rather than by percentage, select the
Absolute intensity errors check box.
16. To specify the threshold spectrum enhancement method as automatic, select the
Automatic intensity threshold check box. This method limits the number of ions in the
final spectrum prior to library searching by applying an intensity threshold. If the
intensity of an ion is below the specified threshold, the ion is discarded from the
spectrum. To set a value for the intensity threshold, use the up and down arrow or type a
value.
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Choosing Report Types
Use the Reports tab to set report values for Survey Screening and Target Matching workflows.
Report values for the Target Screening workflow are set on the Target Screening page, so you
cannot set report values for Target Screening on this page.
Y To specify report types
1. To select a report, click the Reports tab. The Reports Page opens.
2. To generate a particular report, select the Active Report check box next to the report
name. You can also select the Print, Create PDF, and Create XML check boxes as needed
to specify the output type of final report.
• To create the report, select the Active Report check box. You must select this box to
create any type of report.
• To print the report, select the Print check box.
• To create a PDF version of the report, select the Create PDF check box. This report
file resides in the Reports folder in the batch folder.
• To create an XML version of the report, select the Create XML check box. Use this
version to create a record that you can import into a LIMS or electrical LIMS
notebook. This report file resides in the Reports folder in the batch folder.
3. To select a printer for the method, select a printer from the printer list. If you do not
select a printer after selecting the Print check box, Pathfinder alerts you when you try to
save the method.
Do not choose a printer driver that requires user entry into a dialog box before generating
electronic files. An example of this type of printer is the Microsoft XPS Document Writer.
4. In the Method Development area, click Save Method to save the method and make it
available for users.
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Defining Target Screening Settings
Y To set values for Target Screening
1. Click the MS Processing tab. The Target Screening page opens.
2. Browse to select a .csv file as the target database. To edit this file, click Edit. The file
opens in Excel and you can add or change values.
3. To get a report, choose from the listed options. Select Summary report: Create PDF to
get a one page report of the processing results, select Long report: Create PDF to get a
detailed report of reprocessing. You can select to print summary and long reports.
4. To specify a local printer that Pathfinder uses for printing reports, select from the printer
list. If you do not select a printer, Pathfinder has no specified font size. The report might
have small text.
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5. Click Setup to set parameters for target screening processing. The Setup dialog box
opens. For parameter definitions, see “Target Screening Setup Dialog Box” on page 40.
Once you define the screening method as Based on Exactive screening method, set other
available options.
6. Click OK to save Setup values and close the Setup dialog box.
7. Click Save to save the method. The Save Method box opens. Name the method and click
OK.
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Target Screening Setup Dialog Box
Parameter
Description
Screening Method
Select Based on Exactive screening method to set values for
Target Screening.
Company name
Specify the name of company.
Laboratory name
Specify the name of laboratory.
Company logo
Specify the location of the logo file for the company. For
example: C:\logo.jpg.
RT Window (min)
Set the size of the RT window in minutes.
Exact Mass Window
(ppm)
Specify the exact mass variability window. This is the value
above and below the center m/z value.
Exactive
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Adduct 1, 2 or 3
Type a value for Adduct 1, Adduct 2, and Adduct 3, respectively.
For example, H+, NH+, and Na+. The values entered identify
the adducts listed in and applied globally throughout the
configuration file. Adducts are polarity sensitive. For negative
ionization, enter negative adducts.
No Specified Retention
Time
Select the No Specified Retention Time check box if you do
not wish to have retention time specified. Selecting this option
allows Pathfinder to search the entire raw file for the peak
specified. Select this option when you do not know when the
target compound will elute.
Report retention time
based on
If you have selected the No specified retention time option, then
select the First Peak or Highest Peak option to provide guidance
for Pathfinder. This is necessary in case more than one m/z
match is found in the raw file to guide Pathfinder in selecting a
single peak to use for processing. Select this option when you
know when the target compound will elute.
Report All compounds
Listed in Configuration
File
Select the Report All compounds Listed in Configuration File
check box if you want to have reports for all compounds in the
configuration file whether or not matches are found for them. If
this option is not selected, the default is to report on only those
compounds where matches are found in the raw file. This
option applies to an Exactive experiment only.
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Creating a Processing File for Target Screening
A Target Screening workflow requires experiment- and compound-specific information that
you define in the processing configuration file. Name and save the processing configuration
file as a .csv file before setting up a Target Screening workflow.
Define target screening workflows on the computer managing the instruments. You cannot
define target screening workflows on a remote computer.
Y To define the Pathfinder processing configuration file parameters
1. Open Excel and then open the Target Screening processing configuration file, called
Target Database, for the experiment you want to perform. The template for the Target
Screening workflow is in the C:\Pathfinder\Examples directory.
Use the ExactiveTest_Config.csv file in screening experiments that identify compounds
based on accurate mass of parent ion with adducts, retention time, or both. You can view
the processing configuration file in Figure 2, a sample summary report in Figure 5 on
page 67, and a sample long report in Figure 6 on page 68.
2. Set the compound-specific parameters as shown in Table 2 on page 42. Observe the
following:
• Never add, remove, or rearrange columns or rows above row 4 in a processing
configuration file. Pathfinder cannot process the analysis if any information is
missing or moved.
• When you save a .csv file, you lose any changes you make, such as enlarging or
shrinking columns or rows.
• Always save the edited processing configuration file as a .csv file. You cannot run
Pathfinder using Excel (.xls) files.
3. Save the file with a new name in a location of your choice.
Figure 2 provides an example for the Target Screening processing configuration file
(name.csv).
Figure 2.
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Exactive processing configuration file
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Table 2 describes the compound-specific parameters for the Target Screening workflow.
Table 2. Compound-specific parameters
Parameter
Description
Compound Name
(all experiments)
Name of the compound. If a spectra library is implemented,
this name must match the library compound name.
Compound names are required for all screening experiments.
Chemical Formula (Exactive) Chemical formula of the compound.
Polarity (Exactive)
Positive or negative ionization.
Analyte Type
Specifies a parent, metabolite, or other analyte type, as
appropriate. The first letter of an analyte type name is posted
in the report. This parameter is required for all experiments.
The Code column in the reports refers to the information in
the Analyte Type column: P for Parent, M for Metabolite, or
I for Internal Standard.
Expected RT
Expected retention time. This parameter is required for all
experiments.
Intensity Threshold
For MS/MS, and for MS/MS and MS3 experiments, a search
hit is only reported if the signal intensity for the most intense
or most specific fragment in the MS/MS spectra is equal to
or above this value.
For QED MS/MS experiments, a search hit is only reported
if the intensity of the fragment ion collected in an SRM scan
event is equal to or above this value.
For other types of experiments, a hit is reported if the TIC
signal intensity is equal to or above this value.
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Adduct 1, 2, 3 (Exactive)
Ions attached to parent ion during ionization.
Fragment 1, 2, 3 (Exactive)
Ions in the HCD scan, which the user can choose for
additional information about the compound match.
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3 Working with Methods
Working with Pathfinder Methods
Changing a Pathfinder Method
Use the Method view to review or change a method.
Y To review or change a method
1. In the Method Development mode, click Open method in the Method view. The Open
Method dialog box opens.
2. Select an existing method. The Method view opens, displaying the current parameters.
3. Change parameter values on any page and click Save method in the Method
Development area.
4. To save this modified method under a new name, choose File > Save as…
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Working with Methods
Working with Method Groups
Working with Method Groups
Create method groups that define a set of methods to be used for a specific instrument
configuration or task. Certain methods might require that API sources, HPLC systems, or
HPLC solvents be in place. Once you create a method group, you can activate it for users so
that they can only select a method that works with the current configuration of the
instrument.
When creating groups, remember these concepts:
• A method can exist in more than one group.
• You can only make one group at a time available to users by activating it.
• You do not need to create method groups. If this is the case, Pathfinder displays all
methods to the user.
• You can deactivate methods within a group so that they remain in the group but are not
available to users.
• The user only sees available methods, whether part of a group or not.
• The method group capability can be turned off so that all methods are available.
• The method group filter is only active in the User account. An administrator always sees
all methods.
Creating a Method Group
Y To create a method group
1. From the Method Development view, click Method Groups in the Method
Development area. The Method Groups page opens.
2. To activate the method group capability, select the Enable method group filtering for
users check box.
3. Click New Group. The New Method Group area opens.
4. Type a name for the group in the Name box and click Create. The Method Groups page
displays the new group as the active group.
The Available list box displays all Pathfinder methods that are currently available.
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Working with Method Groups
5. To add methods from the Available box, select a method name and click
method to the Selected box.
to add the
6. When the Selected box displays the list of chosen methods, click Save to save the group
with the methods.
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Working with Methods
Working with Method Groups
Modifying a Method Group
Y To add methods in a group
1. Select the method group in the Active group list.
2. Add methods by selecting the names in the Available box and clicking
.
3. Click Save.
Y To remove a method from a group
1. Select the method group in the Active group list.
2. Select the name of the method in the Selected box and click
to remove the method.
3. Click Save.
Y To deactivate a method from a group
1. Select the method group in the Active group list.
2. Clear the check box next to a method in the Selected area. The user will not see the
method when the methods from the group are displayed.
Y To rename a group
1. Select the method group in the Active group list.
2. Click Rename. The New Method Group dialog box opens.
3. Type a new name for the group in the Name box and click Change to rename the group.
4. Click Save.
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Working with Method Groups
Changing the Active Method Group
Use the following procedure to make a different method group active. The Method
Development area displays the active method group in the method group view.
Y To change the active method group
1. Click Method Group view.
2. If the Enable method group filtering for users box is not selected, select it.
3. Select the method group name in the Active group list.
4. Close the dialog box. The new method group appears when you submit samples.
Deleting a Method Group
Deleting a method group does not delete the methods in the group. The methods remain
available for other method groups or as individual tests.
Y To delete a method group
1. Select the method group name in the Active group list.
2. Click Delete. Pathfinder requests confirmation of the action.
3. Click Yes in the confirmation box. Pathfinder deletes the method group.
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Working with Samples
Follow these procedures to test methods and perform analyses for both single samples and sets
of multiple samples.
Contents
• Submitting a Single Sample for Analysis
• Submitting Multiple Samples for Analysis Manually
• Creating Files for Importing Lists of Multiple Samples
• Submitting an Imported List of Samples for Analysis
• Viewing Instrument Status During Processing
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Working with Samples
Submitting a Single Sample for Analysis
Submitting a Single Sample for Analysis
Use the User Mode in the User Administration view to analyze a single sample using a
Pathfinder method.
Y To perform an analysis on a single sample
1. Select User Administration and select the type of workflow in the User mode area.
a. To test a survey screening method, click Survey Screening in the User
Administration view to open the Survey Screening page and specify sample
information.
b. To test a target matching method, click Target matching in the User Administration
view to open the Target Matching page and specify sample information and the
molecular weight.
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4 Working with Samples
Submitting a Single Sample for Analysis
c. To test a target screening method, click Target screening in the User Administration
view to open the Survey Screening page and specify sample information.
To clear all sample information and start over, click Clear. To stop the submission process
completely, click Cancel.
2. Type the user name in the User name box.
3. Type the first sample ID in the Sample ID box.
4. Type the autosampler vial position in the Vial position box. If the vial position value is
not in the correct format for your autosampler, Pathfinder displays an error message when
you click Submit. For more information about the autosampler vial position format, refer
to your autosampler documentation.
5. Type the file name with the information for the sample in the File name box. File names
must be unique for each sample. If Pathfinder provides automatic file naming, the File
name box is grayed out. See “Defining a File Naming Convention” on page 94.
6. If you selected Target Matching, type a molecular weight in the Target MW box. Specify
the level of accuracy in the method.
7. Click Go to submit the sample. If the Go button is grayed out, you are missing
information required for the analysis. Place your mouse over the button and Pathfinder
displays a list of needed information.
8. To view results, click Analysis to open the Analysis view. Click Open sequence and select
a results file from the list displayed. For more information about viewing test results, see
“Viewing Workflow Results” on page 59.
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Working with Samples
Submitting Multiple Samples for Analysis Manually
Submitting Multiple Samples for Analysis Manually
Use the User Mode in the User Administration view and the Add option to analyze a set of
samples using a Pathfinder method.
Y To perform an analysis on a group of multiple samples by manually adding entries
1. Click User Administration to open the User Administration view. Select the type of
workflow in the User mode area. The page for that method opens.
2. Type the user name in the User name box.
3. Type the first sample ID in the Sample ID box.
4. Type the autosampler vial position in the Vial position box. If the vial position value is
not in the correct format for your autosampler, Pathfinder displays an error message when
you click Submit. For more information about the autosampler vial position format, refer
to your autosampler documentation.
5. Type the file name with the information for the sample in the File name box. File names
must be unique for each sample. If Pathfinder provides automatic file naming, the File
name box is grayed out. See “Defining a File Naming Convention” on page 94.
6. If you selected Target Matching, type a molecular weight in the Target MW box. Specify
the level of accuracy in the method.
7. Click Add. The sample information is displayed in the table to the right and Pathfinder
clears the user entry parameter boxes.
8. Type information for the rest of the samples, clicking Add after each sample.
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Submitting Multiple Samples for Analysis Manually
9. To edit information for a sample that you have added to the list, click the sample to move
the sample information back to the sample information area. Make changes and click
Update.
To clear all sample information and start over, click Clear. To stop the submission process
completely, click Cancel.
10. To submit all samples, click Go. If the Go button is grayed out, you are missing
information required for the analysis. Place your mouse over the button and Pathfinder
displays a list of needed information.
11. To view results, click Analysis to open the Analysis view. Click Open sequence and select
a results file from the list displayed. For more information about viewing test results, see
“Viewing Workflow Results” on page 59.
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Working with Samples
Creating Files for Importing Lists of Multiple Samples
Creating Files for Importing Lists of Multiple Samples
To enter groups of samples easily, use the templates in
C:\Thermo\Pathfinder\Templates\ImportTemplates. There are three options for entering
data:
• ImportTABTemplate.csv: a tab-delimited template file
• ImportCSVTemplate.csv: a comma-delimited template file
• ImportXMLTemplate.xml: an easily editable xml file
Y To create a file for the analysis of multiple samples
1. Open a template. You can open any of the templates in any type of editor, including
Microsoft Notepad. You can also use Microsoft Excel to make it easy to work with tables
of information.
2. Enter the sample ID, vial number, and file name for each sample in the designated
column for that row. If the vial position value is not in the correct format for your
autosampler, Pathfinder displays an error message when you submit the samples. For
more information about the autosampler vial position format, refer to your autosampler
documentation.
3. If you are doing a target matching test, enter the molecular weight in the TargetMW area.
If you are not doing a target matching test, delete the portion of the template that
requests a molecular weight. The following example is a .csv batch file for a target
matching test.
If Pathfinder provides file naming and you put file names in the batch file, Pathfinder uses
the file names from the batch file. If the batch file has no file names, Pathfinder provides
file names. For more information about file naming, see “Defining a File Naming
Convention” on page 94.
4. Click Save As to name the file and save it in a convenient location. To use this sample list,
see “Submitting an Imported List of Samples for Analysis” on page 55.
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4 Working with Samples
Submitting an Imported List of Samples for Analysis
Submitting an Imported List of Samples for Analysis
Use the User Mode in the User Administration view and the Import option to analyze a set of
samples using a Pathfinder method.
Y To perform analysis using an imported sample list
1. Click User Administration to open the User Administration view. Select the type of
workflow in the User mode area. The page for that method opens.
2. Type a user name in the User name box.
3. Select a specific Pathfinder method from the Method area.
4. Click Import to select a file containing sample information. A browse window opens. For
information about creating the file, see “Creating Files for Importing Lists of Multiple
Samples” on page 54.
5. Select the multiple samples file and click Open.
6. To edit information for a sample that you have added to the list, click the sample to move
the sample information back to the sample information area. Make changes and click
Update.
To clear all sample information and start over, click Clear. To stop the submission process
completely, click Cancel.
7. To submit all samples, click Go. If the Go button is grayed out, you are missing
information required for the analysis. Place your mouse over the button and Pathfinder
displays a list of needed information.
8. To view results, click Analysis to open the Analysis view. Click Open sequence and select
a results file from the list displayed. For more information about viewing test results, see
“Viewing Workflow Results” on page 59.
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Working with Samples
Viewing Instrument Status During Processing
Viewing Instrument Status During Processing
Use the Real Time Status view to monitor sample analysis.
Y To view instrument status
1. After clicking Go to start an analysis, click
view. The Real Time Status view opens.
or click Real Time Status in the Analysis
The Status view displays acquisition status and the data window shows chromatographic
and mass spectral data as it is acquired.
To view device status below the processing status, click the device name. The status page
for that device opens in the lower left area (Status).
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4 Working with Samples
Viewing Instrument Status During Processing
2. Click the Acquisition Queue tab to open the list of submitted samples. When the sample
has finished, the list displays Complete after the sample name.
3. To remove a sample or sequence from the queue, select the check box next to the name.
Right-click the check box and choose Remove checked from the shortcut menu.
Pathfinder removes the sequence item.
You can only delete an entire sequence before the instrument begins to acquire data in
that sequence. Also, if you submit a sequence of samples, you can only delete samples
before the instrument begins to acquire data from those samples.
The following buttons at the top of the Real time status view help you control
acquisition.
Button
Action
Go
Restarts acquisition from a pause.
Stop
Stops acquisition for the sample in progress and pauses the
acquisition queue. If you click Go, Pathfinder continues with the
next sample in the queue.
Pause
Pauses the acquisition queue. If Pathfinder is currently acquiring a
sample, it will continue acquiring. If you click Go, Pathfinder
continues acquisition on the next sample.
Note You can get a quick look at the system status at any time by looking in the lower
right corner of the Pathfinder window. This area displays a green or red LED and a
description of the number of samples in the queue (if any).
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Viewing Workflow Results
This chapter describes viewing the results of processing a sample or samples using a method.
The results appear on the Data Review page and the Report page. Before viewing data, process
your sample following the instructions in “Working with Samples” on page 49.
Contents
• Viewing Data Results
• Viewing Reports
• Modifying Sample Results
• Reprocessing Samples
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Viewing Workflow Results
Viewing Data Results
Viewing Data Results
This section provides information about viewing the results of a sample after processing.
Pathfinder provides sample details at a variety of levels.
• Viewing Results for a Workflow
• Viewing Peak Details
• Viewing Specific Details
Viewing Results for a Workflow
You can view data results for Target Matching or Survey Screening workflows in the Data
Review area. When viewing these results, you can zoom in on a spectrum by placing the
mouse at the outside edge of the area you want to see in greater detail and sliding the mouse
over the area. If you zoom a full scan spectrum, Pathfinder retains the zoom frame when you
select different peaks in the navigational chromatogram. When you switch from sample to
sample, zoom resets to defaults. When you switch filters within a sample, the zoom resets to
defaults. When you switch to another view and return, Pathfinder maintains the zoom setting.
To view reports from the Target screening workflow, go to the Thermo\Pathfinder\Batches
directory. Open the batch folder and open the Reports folder. Double click to open the .pdf
file.
Y To view data results
1. From the Administrator view, click Analysis. The Analysis view opens.
2. Click Open sequence. The Open sequence dialog box opens. The most recently acquired
sequence is at the top of the dialog box.
3. To select a data results file, select the file name and click Open. The Data Review page
opens, displaying workflow data.
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Viewing Workflow Results
Viewing Data Results
a. If your method was Survey Screening, the Survey Screening page opens.
Figure 3.
Survey screening data review
Sample list
Peak list
Navigational
chromatogram
Integrated peak
chromatogram
Spectrum
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Viewing Workflow Results
Viewing Data Results
b. To view Target matching values, click the Target Matching tab. The Target
Matching page opens.
Figure 4.
Target matching data review
Peak List
Sample list
Navigational
chromatogram
Integrated peak
chromatogram
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Elemental
Composition results
Active data source
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Viewing Data Results
If the report is for Target matching, the Peaks box identifies in blue any peaks that
satisfy the matching criteria. To make sure Pathfinder has identified a peak, hover
over the peak with a mouse. Pathfinder highlights the peak in the Peak list if the peak
has been identified.
On the Data Review page for both Target Matching and Survey Screening, you can
view the results of using a supplemental detector to define the data field by clicking
the arrow in the Data source list.
For a detector type of Analog, A/D, or UV detector channel, Pathfinder stores only the
peak list. For a detector type of PDA, Pathfinder stores only the peak list, but you can
view a PDA spectrum. You cannot perform any other processes (for example, target
matching, target screening, or elemental composition) from any of these supplemental
detectors because they do not have a mass data spectrum.
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Viewing Workflow Results
Viewing Data Results
Viewing Peak Details
Y To view peak information
1. To view peak information, click a specific peak number. Pathfinder identifies the peak in
the navigational chromatogram using a red line that marks the retention time.
Navigational
chromatogram
Integrated peak
chromatogram
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Viewing Data Results
2. To view a different part of the peak and review the spectrum for that area, click an area in
the integrated peak chromatogram.
The retention time marker in the integrated peak chromatogram moves to the area you
clicked and the red line in the navigational chromatogram moves with it.
The Spectrum box displays spectrum information for the selected retention time.
To reset the peak, select another peak in the Peak list and then select the original one.
Viewing Specific Details
If your sample contains HCD data, Pathfinder displays an HCD spectrum in the data review.
The HCD spectrum displayed should be one scan number greater than the full scan of the
target match. Use the HCD scan only to recognize fragments.
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Viewing Workflow Results
Viewing Data Results
Click the Elemental Composition tab to view the values that meet the window of the
elemental composition parameters. The Elemental Composition page opens. The peak with
the best match has a checkmark in the Index box according to the elemental composition
criteria. Pathfinder sorts the peaks by the spectral fit value, from high to low.
You can select a different peak from the Elemental Composition list. To change the best
match choice, select the check box of the compound that is more accurate. Elemental
composition results are also available in the Survey Screening and Target Matching reports.
For Target Matching results, click the Target Matching tab. The Target Matching view shows
detailed information about the selected peak.
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Viewing Workflow Results
Viewing Reports
Viewing Reports
Y To view the test report for Target Screening workflows
1. To view reports from the Target screening workflow, go to the
Thermo\Pathfinder\Batches directory.
2. Open the batch folder and open the Reports folder.
3. Double-click to open the .pdf file.
4. From the .pdf file, choose File > Print to print the file. Even if you did not select a type of
file, Pathfinder creates a .pdf file.
Figure 5.
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Viewing Reports
Figure 6.
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Target Screening sample long report
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Viewing Reports
5. Target screening reports display the following information so that you can review the
data.
For each processed data source:
• The summary report shows
• Header information
• A trace for each target compound
• A table showing matches to the selected database
• The long report shows
• One page for each compound
• Detailed results of all adducts and fragments identified using the database
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Viewing Workflow Results
Viewing Reports
Y To view the test report for Target Matching or Survey Screening workflows
1. In the Analysis view, click Reports view. The Reports page opens.
2. Select a report in the Reports box. Select a specific file in the Sample file box.
3. All reports display the following information so that you can verify the data:
• A batch header at the top with the batch name, the Pathfinder method, the method’s
lab and assay name, and the user who submitted it
• A sample header with sample specific information, such as the sample ID, the date
and time of acquisition, the raw data file name, and the vial position
Figure 7.
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Survey screening report
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Viewing Reports
Survey screening reports display the following information so that you can review the
data. For each processed data source, Pathfinder displays a chromatogram followed by a
report block for each peak detected in that data source, including
• Textual peak information, such as peak retention, height, and area
• An enlarged view of the detected peak (the same data that is in Data Review)
• A mass spectrum taken from the peak apex
• For HCD processing, an HCD trace if one was acquired
The HCD scan shown is one scan number greater than the mass spectrum taken
from the peak apex.
• Elemental composition results
This report block repeats once for each data source that was selected on the Peak
Processing tab of the Method view.
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Viewing Reports
Figure 8.
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Target matching report
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Viewing Workflow Results
Viewing Reports
Target matching reports display the following information so that you can review the data
for each processed data source:
• A table listing all peaks, with a column marked Match to indicate the peak that
matches the user-entered target, if there is one, and a measure of difference
• Textual peak information, such as peak retention, height, and area
• An enlarged view of the detected peak (the same data that is in Data Review)
• A mass spectrum, taken from the peak apex
• For HCD processing, an HCD trace if one was acquired
The HCD scan shown is one scan number greater than the mass spectrum taken
from the peak apex.
4. To export a report as a document, click the Export Report button in the report toolbar.
You can also use the toolbar to go to a specific page of the report and zoom in or out for
viewing report detail.
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Viewing Workflow Results
Modifying Sample Results
Modifying Sample Results
Following automated analysis, use Pathfinder’s Data Review workspace to review the results of
the analysis and make any relevant changes. You can modify samples to change elemental
composition parameters or peak selection. In the Data Review area, you can manually
integrate peaks, add or remove chromatographic peaks, reprocess Target Matching results, and
batch reprocess an entire result set.
CAUTION If another administrator is making changes to a sample, such as reprocessing,
Pathfinder advises you that this action is happening. You might not be viewing an updated
file. While it is possible for two administrators to view the same sample and make changes
to it, such as adding or removing peaks, you might not be making changes to the latest
version of the file.
• Working with peaks
• Changing Parameter Values for a Single Sample
• Reprocessing a Group of Samples
Working with peaks
After viewing the results of the automated processing, you might need to modify some of the
peaks because integration parameters did not adequately process the selected peak, the
selected peak is of no interest, or Pathfinder did not process a peak of interest.
For target matching, Pathfinder identifies and displays only matches within the mass tolerance
and the intensity threshold. If you manually integrate a peak, and it does not meet the method
matching criteria, Pathfinder removes it from the list. If you add a peak and it does not match
criteria, Pathfinder provides a warning message and does not add the peak to the list or the
report.
• To add a peak
• To remove a peak
• To manually integrate a peak
Y To add a peak
1. Right-click the navigational Chromatogram view and choose Add a peak from the
shortcut menu.
2. Click the base line to set the start time for the new peak and drag the cursor to the end
time to define the peak. The chromatogram does not change, but the new peak data is
added to the peak list. The integrated peak shows the peak data.
3. Click the Report tab to view the new peak information.
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Modifying Sample Results
Y To remove a peak
1. Right-click the Integrated peak chromatogram view and choose Remove a peak from the
shortcut menu. Click the area with the peak you want to remove. You can also right-click
the peak in the peak list and choose Remove a peak. A message box opens, requesting
confirmation.
2. Click Yes to remove the identified peak. The chromatogram does not change, but
Pathfinder updates the results set by removing the peak data. Or, click No to leave the
identified peak and select another peak with the cursor.
Y To manually integrate a peak
1. In the lower left corner of the data view, select one of the square blue boxes that define a
peak.
2. Drag the blue box to include any region adjacent to the original peak.
To redraw the peak integration, right-click and choose Rescale to return the peak to its
original state.
Note As an aid to manual peak addition and integration, you can zoom in
horizontally, vertically, or both simultaneously by drawing a line or box on the
Chromatogram plots using a left-click-and-hold mouse movement. To restore the
default plot size, right-click the chromatogram and choose Rescale. You can also
change the spectral plot using the same mouse movements.
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Viewing Workflow Results
Reprocessing Samples
Reprocessing Samples
You can reprocess samples to change peak selection parameters or reporting options. Often
this is done after a preliminary data review. Use the reprocessing options available to make
changes to Pathfinder methods, such as changing peak integration and selection settings. You
can also use those options to create new result sets or to manually modify the target molecular
weight used for Target Matching.
• Changing Parameter Values for a Single Sample
• Reprocessing a Group of Samples
Changing Parameter Values for a Single Sample
Y To change the value for a target match to look for other compounds
1. In the Data Review area, select Open sequence to display a browse box.
2. Select the sequence you want to change. The sequence review opens.
3. In the Target MW column, click the value and type a new number in the box.
4. To process the sample using the changed value, select Enter.
The Peak list identifies peaks that match the new value in blue, but the chromatogram
does not change.
5. To view the report with changed values, in the Analysis view, click Report view to display
a report view. In the report view, click Report Type and select Target Matching from the
dropdown list. Pathfinder generates and opens the new report.
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Reprocessing Samples
Reprocessing a Group of Samples
To reprocess samples using a different method, choose the Reprocess sequence command. If
you want to restore the results from your method after the sequence has been changed
manually, for example, if it contains samples with one or more manual integrations or peak
add/remove operations after the original method processing, you can reprocess the samples
using the original method.
Y To change the processing values for a group of samples
1. To reprocess a sequence, you can change the original processing method or use a different
method. To change a method or create a new method, see “Working with Pathfinder
Methods” on page 21. You can also use the same method to restore original processing
values.
2. To reprocess a sequence, select the Reprocess sequence command in the Data Review
area.
If you do not have a sequence open, a browse box opens. Select a sequence to reprocess.
The Reprocess Sequence dialog box opens displaying the current processing method for
the sequence.
To select a new method, click Browse to open a list of available methods and select a
method for reprocessing.
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Viewing Workflow Results
Reprocessing Samples
3. To generate the reports using an updated or new method, select the Generate reports
check box. If you want to postpone generating reports, you can clear the box, but
Pathfinder deletes all earlier reports in either case.
4. Click Reprocess to review the samples and method in Pathfinder. Pathfinder displays the
results in the Data view after reprocessing the sequence.
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Parameter
Definition
Current method
View the original method Pathfinder used to generate
sequence results. This box cannot be changed.
New method
Select a new processing method.
Generate reports
Generate reports while processing. If you do not select
this option, you can still generate reports by selecting the
Report view command in the Analysis view.
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Managing Users and the Pathfinder System
This chapter describes how to create and modify methods.
Contents
• Working with Users
• Defining System Information
• Troubleshooting
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Managing Users and the Pathfinder System
Working with Users
Working with Users
Use the User Administration view to add, modify, or delete users.
• Adding Users
• Changing User Information
• Removing a User
Adding Users
Use this page to define users who can receive e-mails from the system.
Y To add a user
1. In the Method Development view, click User Administration. The User View opens.
The following fields are not used in Pathfinder.
• Password
• Full name
• Phone number
• Account number
2. Type the user name in the User name box. Enter only alphanumeric characters.
3. Type the user’s system e-mail address in the E-mail address box.
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Working with Users
4. Specify the type of notification the user will receive when a sample submitted by that user
is completed. For information about setting up a virtual e-mail server, see “Setting Up a
Virtual E-Mail Server” on page 85. When doing this task, you might need help from your
IT professional.
• To send no e-mail, select None.
• To send e-mail notification of completion, select E-mail. You can send e-mail for all
types of workflows.
• To send e-mail notification of completion and a report copy, select E-mail and
Report. Pathfinder sends reports from all workflows.
5. Click Add to add the user to the user list.
• To make the user active on the system, select the check box next to the user name in
the Available users area.
• To make the user inactive on the system, clear the check box next to the user name in
the Available users area.
Changing User Information
Y To change user information
1. In the Method Development view, click User Administration. The User View opens.
2. Select the user name that you want to change in the Available users area.
3. Change field data as required. To clear all fields, click Clear.
4. Click Update to save the changes.
Removing a User
Y To remove a user from the Pathfinder system
1. In the Method Development view, click User Administration. The User View opens.
2. Select the check box for the user name that you want removed in the Available users area.
3. Click Delete to remove the user.
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Defining System Information
Defining System Information
Use the Tools menu commands to define certain options used throughout Pathfinder and to
call up the monitoring dialog box for the Pathfinder server.
• Configuring Pathfinder on Remote Workstations
• Setting Up a Virtual E-Mail Server
• Configuring Instruments
• Initializing Pathfinder Using Xcalibur
• Setting Options
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Configuring Pathfinder on Remote Workstations
When you start Pathfinder on the local workstation, both the Web service and the user
interface client start. By default, the user interface client communicates with the Web service
through the routed name localhost, and no further configuration of the client/server
connection is required. The local workstation is automatically ready for use with Pathfinder.
When you start Pathfinder on the remote workstation, the user interface client appears. The
first time you open Pathfinder on that workstation, you must define the Web service
workstation so that the client can communicate with the server.
Y To configure Pathfinder
1. Double-click the Pathfinder icon to open the user interface client.
If this is the first time you have started Pathfinder on the remote workstation, you must
define the Web service workstation in order for the client to begin communicating with
the server.
The Connect to Pathfinder server dialog box initially shows the default server name
localhost.
2. To connect the remote computer to the computer with the main Pathfinder application,
replace localhost with the network PC name of the workstation that is running the Web
service (the local on-instrument installation). Click Test to confirm the connection.
If Pathfinder can connect successfully, it displays a message.
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If Pathfinder cannot connect successfully, it displays a message for any of the following
problems:
• The network is not available and so no connection is possible.
• The network is available, but a computer with that name is not found.
• Pathfinder found the computer, but the Pathfinder application is not installed or is
not installed correctly.
3. After you have designated the new Pathfinder server and the test connection has been
successful, click Apply to save this information for the remote workstation. The remote
workstation is now ready for use with Pathfinder.
If the main Pathfinder computer is not available, Pathfinder displays the following
message: System is not available.
Y To change the target workstation
1. To open the configuration dialog box, type config in the Administrator password field
and click the Login button.
2. Type the name of a new target workstation to change the target instrument computer and
have Pathfinder validate it.
You can change the target workstation at any time.
IMPORTANT Before using Pathfinder to acquire data from your instrument, process one
sample through Xcalibur. For more information, see “Initializing Pathfinder Using
Xcalibur” on page 92.
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Setting Up a Virtual E-Mail Server
Pathfinder users can register to receive e-mail notifications regarding the samples submitted
using their name. There are two types of e-mail notifications:
• An e-mail notification stating that the sample has completed
• An e-mail notification stating that the sample has completed with a report attached.
Pathfinder sends an e-mail for each sample as it completes. There is no e-mail sent when a
sequence of multiple samples completes.
Pathfinder uses the Default SMTP Virtual Server that is installed with Microsoft Internet
Information Services. You should install Pathfinder on the instrument behind a firewall and
on the same domain where you have defined the users who receive notifications. Please work
with your IT department to make sure these changes are compatible with your e-mail server.
This section provides top level instructions for setting up a virtual e-mail server using the
Default SMTP Virtual Server application. For more information, refer to your computer
documentation or your IT professional.
Y To set up a virtual e-mail server
1. From the computer that will send e-mail notification, choose Start > Control Panel >
Administrative Tools > Internet Information Services. The Internet Information
services view opens.
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2. Right-click Default SMTP Virtual Server and choose Properties. The Default SMTP
Virtual Server Properties dialog box opens.
3. Click the Access tab. The Access page opens. Click Connection. The Connection page
opens.
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4. Select an option, add computers to the list, and click OK.
• For the highest level of security, select the Only the list below option and add only
the computer that you are configuring.
• For more general access, select the All except the list below option. This option
allows you to include all computers except those you add to the list.
When you click Add, the Computer dialog box opens. After adding computers, click OK.
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5. In the Default SMTP Virtual Server Properties dialog box, click Relay. The Relay
Restrictions dialog box opens.
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6. Select an option, add computers to the list, and click OK.
• For the highest level of security, select the Only the list below option and add only
the computer that you are configuring.
• For more general access, select the All except the list below option. This option
allows you to include any computer except those listed in the dialog box.
7. To add operator permission for local computer/ASPNET, click the Security tab. The
Security page opens.
localcomputer\ASPNET
8. Click OK.
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9. Click Add to select local computer\ASPNET and click OK. The application adds the
name to the security list.
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10. Log into Pathfinder as an administrator and select Tools/Options. The Options dialog
box opens. Click the Notification tab and make sure the SMTP server name is the same
as the one on the Security page. Click OK to set up notification. If you did not specify a
computer on the Security page, type localhost for the SMTP server name.
localhost
Configuring Instruments
To use instruments with Pathfinder, you must define and configure those instruments.
Choose from one of the following procedures to configure instruments or determine
instrument status:
• Adding Hardware Devices to the Pathfinder Configuration
• Removing Hardware Devices from the Pathfinder Configuration
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Adding Hardware Devices to the Pathfinder Configuration
Y To add hardware devices to Pathfinder
1. To make changes to your hardware configuration, close Pathfinder and Xcalibur, if
opened.
2. Choose Start > Programs > Thermo Foundation 1.0 > Instrument Configuration.
The Instrument Configuration window opens.
3. From the Device Types list, select the type of hardware device to add to your instrument
configuration. The Instrument Configuration window displays all available configurable
devices as buttons in the Available Devices area.
4. Click the button of the device you want to add. The button depresses to indicate that it is
selected.
If you do not see the device you want to configure, you might need to install the device
driver.
5. To add the device to the Configured Devices area, click Add. The software copies the
selected Available Devices button to the Configured Devices area, displayed as a
Configured Devices button.
To quickly add a device from the left column, double-click the device button.
6. To select a device to configure, click the device button in the Configured Devices area.
The Configured Devices button depresses to indicate that it is selected.
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7. To configure the selected device, click Configure. The DeviceName Configuration dialog
box opens (DeviceName corresponds to the selected device).
To quickly configure a device in the right column, double-click the device button.
8. Enter all required configuration information for the device. Complete entries and options
for all pages. Options on each page are different for every device.
9. To save settings and close the DeviceName Configuration dialog box, click OK. The
Instrument Configuration window stays open.
10. Repeat steps 3 through 9 for all devices to be configured.
11. To save settings and close the Instrument Configuration window, click Done.
Once you have added all hardware devices to the system, see “Creating an Instrument
Method” on page 19 for information about setting up an instrument method to work
with Pathfinder.
IMPORTANT Before using Pathfinder to acquire data from your instrument, process one
sample through Xcalibur. For more information, see “Initializing Pathfinder Using
Xcalibur” on page 92.
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Removing Hardware Devices from the Pathfinder Configuration
Y To remove hardware devices from Pathfinder
1. To make hardware changes effective, close Pathfinder and Xcalibur, if opened.
2. Choose Start > Programs > Thermo Foundation 1.0 > Instrument Configuration.
The Instrument Configuration window opens.
3. To select a device to remove from the configuration, click the device button in the
Configured Devices area. The Configured Devices button depresses to indicate that it is
selected.
4. To remove the selected device, click Remove. Pathfinder removes the selected Configured
Device button from the Configured Devices area.
5. Repeat steps 2 through 4 for all devices to be removed.
6. To save settings and close the window, click Done.
Initializing Pathfinder Using Xcalibur
Before you process a sample using a new Pathfinder installation, process a sample through
Xcalibur using a configured instrument to set the vial positions for that instrument.
Before using this procedure, install all software including Pathfinder and configure the
instruments.
• For information about installing your software, refer to the CD insert.
• For information about configuring Pathfinder, see “Configuring Pathfinder on Remote
Workstations” on page 83.
• For more information about configuring instruments, see “Configuring Instruments” on
page 89.
Y To process a sample using Xcalibur
1. To open Xcalibur, double-click the Xcalibur icon on your desktop (
Start > All Programs > Xcalibur.
) or choose
2. Following the instructions in the Help using the “Working with Processes and Sequences”
section (or the Thermo Xcalibur Acquisition and Processing User Guide), run a sample
through your configured instrument.
IMPORTANT Each time you configure a new instrument, you must run a single
sample through Xcalibur to correctly set the tray configuration used during vial
position validation.
After running the Xcalibur sample, use Pathfinder for acquisition.
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Setting Options
Set options from the Tools menu to define display characteristics, file naming convention,
system availability and status, and notification capabilities for Pathfinder.
• Setting Display Options
• Defining a File Naming Convention
• Limiting System Access
• Specifying Notification
Setting Display Options
Use the Display tab in the Options dialog box to define the display precision (the number of
decimal points) used throughout Pathfinder.
Y To set display options
3. Choose Tools > Options. The Display dialog box opens.
4. Click the Display tab. The Display page opens.
5. Enter the number of decimal places to be displayed for retention times in the Retention
times box.
6. Enter the number of decimal places to be displayed in m\z values in the m\z values box.
7. Enter the number of decimal places to be displayed for peak responses (height or area) in
the Responses box.
8. Under Graphics, enter a value in minutes to specify the width of the integration peak
window in the Peak window box.
9. Click OK to apply your format choices and close the Options dialog box. Or to apply the
changes to the current display and keep the dialog box open, click Apply. To close the
dialog box without making changes, click Cancel.
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Parameter
Definition
Decimal precision
Retention times
Specify the number of decimal places to be displayed for
retention times in a report.
m\z values
Specify the number of decimal places to be displayed for m\z
values in a report.
Responses
Specify the number of decimal places to be displayed for
responses in a report.
Graphics
Peak window (in minutes)
Specify the time for the peak window in minutes.
Defining a File Naming Convention
Use the File Naming tab in the Options dialog box to activate automatic RAW file naming
and to define the file naming system provided by Pathfinder for a user submission.
Pathfinder provides the ability to specify a naming convention to use to name files. This
function helps create consistency among data files and eliminates manual entry errors for file
names. You can design file names using:
• Time stamp (The time the test was performed)
• Sample ID
• Index
• Vial position
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Y To select a file naming convention
1. Choose Tools > Options. The Options dialog box opens.
2. Click the File Naming tab. The File Naming page opens.
3. Select the Enable automatic file naming check box to turn on this feature. To turn off
automatic file naming, clear the Enable automatic file naming check box.
4. Select an option from the File name format list.
5. Click OK to apply your format choices and close the Options dialog box. Or to apply the
changes to the current display and keep the dialog box open, click Apply. To close the
dialog box without making changes, click Cancel.
Parameter
Definition
Naming options
Enable automatic file naming
Turn on automatic file naming.
File name format
Select a format to organize files by name. You can sort files by
the sample ID or date and time of acquisition.
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Limiting System Access
Pathfinder provides the ability to limit system access. Select from either open access
(administrator and user availability) or restricted access (administrator-only use). Use the
System tab in the Options dialog box to set the system for Administrator-only use. You can
also indicate the state the instrument should move to after acquisition is complete.
Y To limit system access
1. Choose Tools > Options. The Display dialog box opens.
2. Click the System tab. The System page opens.
3. In the Availability list, select either Open Access to allow users and administrators to use
the system, or Restricted Access to allow only administrators to use the system. In
restricted mode, the user client indicates that the system is not available and displays no
workflow buttons.
4. Click OK to apply your format choices and close the Options dialog box. Or to apply the
changes to the current display and keep the dialog box open, click Apply. To close the
dialog box without making changes, click Cancel.
Parameter
Definition
Availability
Define system access. Select from these options:
• Open Access: To allow users and administrators to use the system.
• Restricted Access: To allow only administrators to use the system.
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Specifying Notification
Use the Notification tab in the Options dialog box to define the e-mail notification options to
be used and to supply credentials for certain security requirements for e-mail activities.
Pathfinder sends notification and reports for all workflows.
When you specify a notification method in the Notification allowed box in the user view,
Pathfinder sends an e-mail when a sample is finished. To determine the sender name and user
values, set them on the Notification page. For more information about setting up an e-mail
server, see “Setting Up a Virtual E-Mail Server” on page 85.
Y To set notification parameters
1. Choose Tools > Options to open the Options dialog box. Click the Notification tab.
The Notification page opens.
2. Set the values in the window and click Apply.
Parameter
Definition
SMTP server name
Identify the server name where Pathfinder resides. For more information about
identifying or setting up an SMTP server, see “Setting Up a Virtual E-Mail Server” on
page 85 or contact your IT professional.
SMTP port
Specify the port.
Sender’s email address
Change the sender e-mail address.
CC sender
Copy the sender with this notification.
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Troubleshooting
Troubleshooting
The following list of best practices and troubleshooting techniques can provide help when you
receive unexpected results from a sample or your system seems to have problems.
To see if the server is connected
Type config for the Admin password to open the following dialog box. Click Test
to see if a server is connected.
If you get a “Pathfinder is offline” Confirm that both the remote and instrument PCs are connected to the network
message
Confirm that both PCs are on the same Windows domain.
Confirm that the correct computer name was entered for the Instrument PC. To
find the computer name:
On the Instrument PC, right-click on My Computer and select Properties |
Computer Name tab
Your system lists the full computer name and domain. Pathfinder only
requires the portion of the computer name before the domain. For example if
the full computer name is JOHN-DOE-PC.region.company.com Pathfinder
only requires “JOHN-DOE-PC”
Verify that there are methods defined in Pathfinder.
Check Real time status to see if device status is updated.
Check that Pathfinder matches the Xcalibur device status.
If reports weren’t generated
Confirm that the method has Active Reports selected along with one of the other
selections.
If notifications weren’t received
Confirm that the name on the batch matches a user in the system, that the e-mail
address for that person is defined and correct, and that mail notifications were
enabled.
If you don’t see reports when you Reprocess the sequence using the same method file.
open a sequence
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Pathfinder Reference
This appendix describes all the Pathfinder views, commands, and options.
Contents
• Pathfinder Menus
• Method Development View
• Analysis View
• User Administration View
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Pathfinder Menus
Pathfinder Menus
See these sections for a definition of menu options.
• File Menu
• Tools Menu
• Go Menu
File Menu
This section describes all File menu commands in Pathfinder. Different views display different
menu commands, but this list defines all of them.
Command
Description
New
Open a new file.
Open
Open a sample file.
Save
Save a sample file.
Save As
Save a file under a different name.
Print
Print a report.
Print Preview
Check a report before printing.
Log Off
Close Administrator views.
Command
Description
Options
Open the Display options views.
Tools Menu
Set these options:
• Display
• File naming
• System
• Notification
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Go Menu
Command
Description
Method View
Open the Method view.
Instrument Method View
Open the Instrument method view.
Method Group View
Open the Method group view.
Real Time Status
Open the Real time status view.
Data Review
Open the Data review view.
Report View
Open the Report view.
User Mode
Open the User mode view.
User View
Open the User view.
Command
Description
Contents
View the entire Help.
Index
View the index for Help.
Search
View the search function for Help contents.
About
View information about Pathfinder.
Help Menu
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Method Development View
Method Development View
• Method View
• Instrument Configuration Views
• Method Group Page
Method View
Use the Method view to create and distribute a new method or update a method. Users run
samples in the User mode using these methods.
Create a new method in the Method view by using the tabbed pages to define your method
criteria. Use the default settings to create a quick method or customize your own method to
include more detailed method criteria. For more information about using this view, see
“Creating a Pathfinder Method” on page 21 or “Changing a Pathfinder Method” on page 43.
• General Page
• Peak Processing Page
• MS Processing: Elemental Composition Page
• Reports Page
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Method Development View
General Page
Use the General page to set and define the criteria for a new method for all workflows. All
fields must be filled in with the exception of the description. For more information about
using this page, see “Defining General Values for a Method” on page 22.
Parameters
Description
Method name
Specify the unique name of the method. The name you type is visible in the User mode.
Lab name
Define the name of the lab processing the data.
Assay type
Define the type of test.
Injection volume
Define the injection volume in microliters of sample to be injected. Click the up or down
arrows to change the value.
Valid values: 0.1 to 100 μL
Default: 10
Instrument method
Select the instrument method used for data acquisition.
Workflow
Select from available workflows:
• To perform overall peak processing and elemental composition (for the mass
spectrometer data), select Survey Screening.
• To perform overall peak processing and elemental composition (for the mass
spectrometer data), and then compare each MS-detected peak to a user-supplied target
molecular weight to find a target compound in a sample, select Target Matching.
• To search for a targeted list of compounds from a database, select Target Screening.
Description
Thermo Scientific
Include additional information about the new method. This information is visible and
accessible in the User mode and provides key details of the particular method.
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Peak Processing Page
Use the Peak processing page to define the data sources and peak criteria for your Survey
Screening or Target Match method. You can control the peak threshold, limit the retention
time range, and define the sensitivity settings.
• Processing Page
• MS, PDA, UV, and A/D Interface Peak Detection Pages
Processing Page
Use the Processing page to specify data sources and make targeting adjustments for a Survey
Screening or Target Match method. For more information about using this page, see “Setting
Peak Processing Values” on page 24.
Parameters
Description
Data source
MS
Select trace types for mass spectrometric data. MS results provide a peak list, elemental
composition (if requested), and specialized processing for the specific workflow.
Mass range
Process the mass range chromatogram. This feature is only available with Target
Matching.
TIC
Process the total ion current [TIC] chromatogram.
Base peak
Process the base peak chromatogram.
PDA
Select trace types for PDA detectors. For PDA data, valid trace types are Wavelength
range, Total scan, or Spectrum max. PDA results provide a peak list and a viewable
PDA spectrum.
Total scan
Process the entire scan.
Spectrum max
Process the wavelength range for the Spectrum max.
Wavelength range
Process a wavelength range.
UV channel
Select UV detector data. For UV detector data, Pathfinder supports four channels
(labeled Channel A-D). UV results only provide a peak list because no mass or optical
spectra are available.
A/D interface
Select A/D card data. For data from an A/D Card, Pathfinder supports two channels
(labeled A/D Card Ch 1-2). A/D results only provide a peak list because no mass or
optical spectra are available.
Delay offsets
Define an offset of supplemental detector data relative to the MS data.
PDA offset (min)
Set the offset for PDA data.
UV offset (min)
Set the offset for UV data.
A/D offset (min)
Set the offset for A/D data.
Target matching parameters
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Parameters
Description
Positive ion adducts
Select positive ion adducts.
Negative ion adducts
Select negative ion adducts.
Custom
Specify a value to adjust for a system that produces a different adduct.
Valid values: –100 to 100
Default: 0
Mass window (+/-)
Define the mass window size. The default is a search window of 5 ppm. If you use the
default, Pathfinder searches a window that is 5ppm above the target match value and
5ppm below it.
Units
Select the unit type for the mass window.
Minimum acceptable relative Set a value for the minimum acceptable relative intensity by entering a percentage
intensity (%)
value.
Valid values: 1 to 100%
Default: 10%
MS, PDA, UV, and A/D Interface Peak Detection Pages
Use these Peak Detection pages to define parameters for processing samples for a Survey
Screening or Target Match method. For more information about using this page, see
“Defining Peak Detecting and Integration Values” on page 29.
Parameter
Description
Sensitivity
Define the sensitivity settings by selecting Standard (Genesis) or High (ICIS). Once you
select an option, an area opens and provides an opportunity to specify settings for that
option.
• ICIS Peak Integration Area
• Genesis Peak Detection Area
Default: High (ICIS)
Limit the retention
time range
Define the retention time range of the data you want to process and report.
Valid values: 0 to 999
Default: 0 to 999 (the entire analysis)
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Parameter
Description
Enable peak threshold
Define the minimum height a chromatographic peak must achieve to be reported. Define
whether you want to process and evaluate the peak using peak height or area. The
minimum height is a percentage of the largest peak.
Valid values: 0 to 100
Default: 0 (all peaks)
Only select top peaks
Specify that only significant or the largest chromatographic peaks are processed and
reported. Select the number of peaks to view and whether to evaluate the peaks by height or
area.
Valid values: 200 for MS input and 50 for all other detectors
Default: 1 (at least one peak)
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ICIS Peak Integration Area
Use the ICIS Peak Integration page of the Info Bar to specify peak detection and integration
criteria that Pathfinder applies to the active raw file. For more information about using this
area, see “Setting High (ICIS) Integration Values” on page 30.
Parameter
Description
Smoothing points
Define the number of points Pathfinder uses for chromatogram smoothing. Select
an odd number for the number of smoothing points. The valid range is 1 for no
smoothing, and from 3 for minimum smoothing to 15 for maximum smoothing.
To change the number of smoothing points, type a new number of points in the
Points box. The default value is 1.
Baseline window
Specify the number of scans to review, looking for a local minima. The valid range
is 1 through 500. The default value is 40 scans.
Area noise factor
Specify the noise level multiplier used to determine the peak edge after the location
of the possible peak. The valid multiplier range is 1 through 500. The default
multiplier is 5.
Peak noise factor
Specify the noise level multiplier used to determine the potential peak signal
threshold. The valid multiplier range is 1 through 1000. The default multiplier
is 10.
Constrain peak width
Limit the peak width of a component during peak integration of a chromatogram.
You can then set values that control when peak integration is turned on and off by
specifying a peak height threshold and a tailing factor. To constrain a peak width,
select the Constrain peak width check box and enter values in the Peak height (%)
box and the Tailing factor box.
Peak height (%)
View or change the percent of the total peak height (100 percent) that a signal
needs to be above the baseline before integration is turned on or off. This box is
active only when you select the Constrain peak width check box. The valid range is
0.0 to 100.0%. To enter a height, type the appropriate value in the Peak height box.
The default is 5%.
Tailing factor
View or change a factor that controls how Pathfinder integrates the tail of a peak.
This factor is the maximum ratio of the trailing edge to the leading side of a
constrained peak. This box is active only when you select the Constrain peak width
check box. The valid range is 0.5 through 9.0. The default is 1.
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Genesis Peak Detection Area
Use the Genesis Peak Integration area of the Info Bar to specify peak identification and peak
integration criteria to be applied to the active raw file. For more information about using this
area, see “Setting Standard (Genesis) Integration Values” on page 31.
Parameter
Description
Peak parameters
Smoothing points
Define the number of points Pathfinder uses for chromatogram smoothing. Select an
odd number for the number of smoothing points. The valid range is 1 for no
smoothing, and from 3 for minimum smoothing to 15 for maximum smoothing. To
change the number of smoothing points, type a new number of points in the Points
box. The default value is 1.
S/N threshold
View or change the threshold for detecting peak edges. The default value is 0.5 and the
valid range is 0.0 to 999.0. Pathfinder calculates the signal-to-noise ratio using only
baseline signal. Any extraneous, minor, detected peaks are excluded from the
calculation.
Enable valley detection
Use the Pathfinder valley detection approximation method to detect unresolved peaks.
This method drops a vertical line from the apex of the valley between unresolved peaks
to the baseline. The intersection of the vertical line and the baseline defines the end of
the first peak and the beginning of the second peak. To turn this method on, select the
Valley Detection check box. To turn this method off, clear the check box.
Expected width
View or change the expected peak width parameter (in seconds). This value controls the
minimum width that a peak is expected to have if valley detection is selected.
With valley detection selected, Pathfinder ignores any valley points nearer than expected
width/2 to the top of the peak. If a valley point is found outside the expected peak
width, Pathfinder terminates the peak at that point. Pathfinder always terminates a peak
when the signal reaches the baseline, independent of the value set for the expected peak
width. The valid range is 0.0 to 999.0 seconds. To change the current value, type a new
width in the Expected Width box. The default is 0.
Constrain peak width
Limit the peak width of a component during peak integration of a chromatogram. You
can then set values that control when peak integration is turned on and off by specifying
a peak height threshold and a tailing factor. To limit a peak width, select the Constrain
Peak Width check box.
Peak height (%)
View or change the Peak Height where Pathfinder tests the width of target peaks. You
can enter any value from 0% to 100%. The default value is 5.0%.
Tailing factor
View or change a factor that controls how Pathfinder integrates the tail of a peak. This
factor is the maximum ratio of the trailing edge to the leading side of a constrained
peak. This box is active only when you select the Constrain Peak Width check box. The
valid range is 0.5 through 9.0. The default is 2.
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MS Processing: Elemental Composition Page
Use the Elemental Composition page to set parameters that find a list of the closest
compounds to the base peak. When you process a Survey Screening sample, the base peak is
the largest peak in the sample defined by height. For a Target Matching method, the base peak
is the peak closest to the value entered in the Target MW field. Pathfinder sets the specified
base peak to 100 and defines all other peaks as relative in concentration to that peak. For more
information about using this area, see “Specifying MS Processing Values” on page 32.
Parameters
Description
Common parameters
Charge
View or change the charge state you want Pathfinder to use to calculate probable
formulas. To change the charge state, enter a number from –99 to +99. To calculate
formulas and display them in the Results box, click Calculate.
Valid values: –99 to 99
Default value: 1
Nitrogen rule
Select whether or not to use the Nitrogen Rule in the formula calculation. For an
LC-MS analysis, select Do not use because the sample has an even number of
electrons.
For molecular ions of even or odd molecular weight, specify that formulas contain
either an even or an odd number of nitrogen atoms, respectively, in the Nitrogen-Rule
list. Conversely, for fragment ions of even or odd molecular weight, specify the
reverse—that is, specify odd or even, respectively, in the Nitrogen-Rule list.
McLafferty states the Nitrogen Rule as follows: “If an odd-electron ion contains no (or
an even number of ) nitrogen atoms, its molecular ion will be at an even mass
number...[Similarly,] an odd-electron ion will be at an odd mass number if it contains
an odd number of nitrogen atoms.”
Valid values: Do not use, Odd electrons, Even electrons
Default value: Do not use
Mass tolerance (ppm)
Specify a mass tolerance to restrict the number of possible elemental compositions.
Pathfinder returns results of the elemental composition search only if the computed
formula matches the entered mass within the specified tolerance.
Default value: 10 ppm
Pathfinder uses this value for mass tolerance throughout the application for elemental
composition.
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Parameters
Description
RDB equiv low
View or change the low value for double bonds and ring equivalents—a measure of the
number of unsaturated bonds in a compound—and limit the calculated formulas to
only those that make sense chemically. You can specify limits in a range from –1000.0
to +1000.0.
The value is calculated by the following formula:
imax
∑ Ni ( Vi – 2 )
i
D = 1 + ----------------------------------------2
Where
D is the value for the RDB
imax is the total number of different elements in the composition
Ni is the number of atoms of element i
Vi is the valence of atom i
The calculation results in an exact integer such as 3.0, indicating an odd-electron ion,
or an integer with a remainder of 0.5, indicating an even-electron ion. A value of -0.5
is the minimum value and corresponds to a protonated, saturated compound (for
example, H3O+).
Valid values: –1e+003 to 1000
Default value: –1
This value must be less than RDB equiv high.
RDB equiv high
View or change the high value for double bonds and ring equivalents–a measure of the
number of unsaturated bonds in a compound–and limit the calculated formulas to
only those that make sense chemically. You can specify limits in a range from –1000.0
to +1000.0.
Valid values: –1e+003 to 1000
Default value: 100
This value must be greater than RDB equiv low.
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Parameters
Description
Max results
View or change the maximum number of formulas you want Pathfinder to display. To
increase or decrease the value, enter a number less than 400. To calculate formulas and
display them in the Results box, click Calculate.
Valid values: 1 to 400
Default value: 10
Elements in use
Use representative elements
Select to use representative elements to analyze the sample. Turning on representative
elements feeds the list of naturally occurring isotopes to the periodic table that selects
masses. Turning off representative elements feeds the list of elements as they occur in
proteins to the periodic table.
The default value is selected.
Use
Click to use an element during analysis. When you clear the Use check box for a row,
the row becomes grayed out. You can edit other rows.
The default value is selected. By default, the elements listed are C, H, O, and N.
Element
Chemical abbreviation for the element.
Valid values can be any symbol found in the periodic table. No default applies.
Nominal
Define the atomic mass of the represented element.
The default value is the atomic mass of the represented element.
Minimum
Set the minimum number of isotopes that can be used.
Valid values: 0 to 10000
Default value: 0
Maximum
Set the maximum number of isotopes that can be used.
Valid values: 0 to 10000
Default value: 10
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Parameters
Description
Advanced Parameters
Spectral fit parameters
Expected mass error (ppm)
The Spectral Fit algorithm compares the differences between theoretical and measured
isotope masses and intensities. In order to decide whether two peaks have a good or
bad mass fit, Pathfinder must know the expected mass error of the mass spectrometer.
This parameter specifies a value for the expected mass error in the spectrum data to be
used by the algorithm.
For best results, set the expected mass error to a value that causes less than 98 percent
of all mass errors to be smaller than the Expected Mass Error. Specify the expected
mass error in ppm.
Valid values: 0 to 1000
Default value: 3
Expected intensity error (%)
The Spectral Fit algorithm compares the differences between theoretical and measured
isotope masses and intensities. In order to decide whether two peaks have a good or
bad intensity fit, the algorithm must know the expected intensity error of the mass
spectrometer.
This parameter sets a value for the expected intensity error in the spectrum data to be
used by the algorithm.
For best results, set the expected error to a value that causes less than 98 percent of all
intensity errors to be smaller than the Expected Intensity Error value. Specify the
expected intensity error as a percentage.
Valid values: 0 to 100
Default value: 0.5
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Parameters
Description
Missing peak penalty
Set a penalty to identify peaks that are specifically searched for but not found. If the
theoretical isotope pattern of a candidate and the measured isotope pattern are
compared, Pathfinder might not find a measured packet for a theoretical isotope peak.
This is called a missing peak.
The Spectral Fit algorithm assigns a penalty for each missing peak based on the
signal-to-noise of the missing peak, using a low penalty for a low signal-to-noise ratio
and a high penalty for a high signal-to-noise ratio. This penalty can be low, medium,
high, or automatic.
Valid values:
• Automatic: The penalty applied depends on the signal-to-noise ratio of the
missing peak. For a peak slightly above noise, Pathfinder applies a low penalty, for
a peak well above noise, Pathfinder applies a high penalty, and Pathfinder applies a
medium penalty for all other peaks.
Use the automatic penalty only for Orbitrap or FT spectra because only these
results contain noise values. If you select automatic penalty with other spectra (for
example, Quantum or DFS), a medium penalty is applied.
• 1 std dev: Low penalty scoring
• 4 std dev: Medium penalty scoring
• 16 std dev: High penalty scoring
• Closest match: Scoring based on the closest match
If you select a specific value, Pathfinder performs only one type of penalty scoring. You
have a wider scope for measuring penalties if you select Automatic.
Default value: Automatic
Mass error weighted by
abundance
Decide whether the observed mass error on an isotope peak depends on its abundance.
Mass errors on large peaks are usually smaller than mass errors on small peaks. This,
however, is data (mass spectrometer) dependent.
If you select the MassError is Abundance Weighted option, an observed error on a
10% BPI (base peak intensity) peak is valued at only one tenth of the same error on
the base peak.
If you clear the MassError is Abundance Weighted option, an observed error on a 10%
BPI (base peak intensity) peak counts the same as an equal error on the base peak.
The default value is selected.
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Parameters
Description
Intensity error weighted by
abundance
Use peak abundance to determine the observed intensity error on an isotope peak.
Intensity errors on large peaks are usually smaller than on small peaks. This value is
data (mass spectrometer) dependent.
If you select the Intensity Error is Abundance Weighted option, an observed error on a
10% BPI (base peak intensity) peak is valued at only one tenth of the same error on
the base peak.
If you clear the Intensity Error is Abundance Weighted option, an observed error on a
10% BPI (base peak intensity) peak is valued the same as an equal error on the base
peak.
The default value is selected.
Absolute intensity errors
Specify whether the errors are treated as absolute or relative errors.
For example, if the theoretical intensity of a peak is 10% and the measured intensity is
11%, then the absolute error is 1% whereas the relative error is 10%. The difference
between two errors is the absolute error, whereas the percent difference of the expected
intensity is the relative error.
For best results, select this parameter option.
When you select the Absolute intensity errors option, you cannot make changes in the
Intensity % control. Clear the Absolute intensity errors option to change the
Intensity % edit value.
The default value is selected.
Automatic intensity
threshold
Specify the threshold spectrum enhancement method as automatic. This method
limits the number of ions in the final spectrum prior to library searching by applying
an intensity threshold. If the intensity of an ion is below the specified threshold, the
ion is discarded from the spectrum.
When you turn on absolute intensity errors, the Intensity % control is disabled. The
default value is cleared.
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Parameters
Description
Intensity % threshold
Set a percentage to define which peaks of the isotope pattern are included in
determining the spectral distance. If you select Automatic intensity threshold, this
option is grayed out.
For example, if the intensity threshold is set to 1.0% and the isotope pattern of an
elemental composition candidate has the following peaks:
Mass Intensity [%]
222.0 100.00
223.0 20.48
224.0 3.87
225.0 0.99
then only the three peaks with a relative intensity > 1.0% are used to calculate the
spectral distance. The peak at mass 225.0 with a relative intensity of 0.99% would
only be used if the intensity threshold is set to 0.99 or lower.
If the MS data you are analyzing contains noise values (if the data was acquired with
an FT or Orbitrap analyzer), then you should select the Automatic Intensity
Threshold option.
In this case the intensity threshold is set so that only peaks from the theoretical pattern
that have a S/N > 3 are used in the SD calculation.
If the MS data you are analyzing does not contain noise values (if the data was
acquired with an analyzer other than FT or Orbitrap), you should estimate the noise
and select a threshold appropriately.
If your data does not contain noise information (all spectra not acquired with an
Orbitrap or FT instrument), the intensity threshold is internally set to 1%.
Valid values: 1 to 100%
Default value: 0
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Method Development View
Instrument Configuration Views
Use these views to configure and view status on instruments or remove out-of-date drivers.
• Instrument Configuration Window
• Out Of Date Instrument Drivers Detected Dialog Box
Instrument Configuration Window
Use the Instrument view to define values for running an instrument and evaluating how an
instrument performs a test sample. For information about the instrument parameters and
settings, refer to the documentation that came with your instrument. For more information
about using this window, see “Creating an Instrument Method” on page 19 or “Changing an
Instrument Method” on page 20.
Error View
Parameter
Definition
Crash server in acquisition
Fail methods
Prepare for run
Start run
Stop run
Validate method
Use error logging
XML input grid
Select to fill the input grid with error information
Error Grid Values
Parameter
Description
Time (min)
Date/Time
View when the event occurred.
Dataset Name
View the dataset that contains the affected files.
Computer Name
View the name of the workstation initiating the event.
User Name
View the logon name of the user who initiated the event. The administrator of the network
assigns logon names for each user.
Full Name
View the descriptive name of the user who initiated the event. Often, this is the first and last
name of the user. The administrator of the network assigns a full name to the logon name
for each user.
Application Name
View the name of the software application that is associated with the event.
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Parameter
Description
Event
View what occurred. Typical events include importing and exporting data files, methods,
and sequences.
Response
View the action taken by the user (if any) in response to the event.
Comment
View the comment associated with the event.
Buttons
Sort
Specify how you want to sort the entries in the Audit Viewer.
Filter
Specify how you want to filter the entries in the Audit Viewer.
Auto Width
Expand the grid columns to display the longest entry in that column.
Print
Print an active file or document.
Refresh
Update the log.
Out Of Date Instrument Drivers Detected Dialog Box
This dialog box opens only when you choose Start > Programs > Thermo Foundation 1.0
>Instrument Configuration. The Instrument Configuration window opens. If your software
is out of date, Pathfinder displays the following message:
The device drivers for the following instruments are not compatible with the installed
version of Xcalibur. You must install the current drivers before you can use these
instruments.
If this message appears, install the latest software for the instruments listed. This software is
located on an Xcalibur CD-ROM. Run the appropriate setup file to load the new software.
Run Setup.exe from the appropriate instrument directory on the CD-ROM.
Thermo Scientific
Parameter
Description
Instrument
View currently installed instruments with out-of-date software.
Instruments installed in this context indicate that the Setup.exe
files for the listed instruments have been previously run from the
appropriate directory of an Xcalibur CD-ROM.
In Use
View the status of the instrument displayed in the Instrument list:
In Use (Yes) or Not In Use (blank). In Use devices appear in the
Configured Devices area of the Instrument Configuration
window.
Version
View the current version of the instrument that is displayed in the
same row of the Instrument list. Make sure the version to be
installed is more recent than the current version.
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Analysis View
Method Group Page
Use the Method Group page to set and establish settings for a specific device. You can create a
new group or update and modify an established group setting. For more information about
using this page, see “Creating a Method Group” on page 44, “Modifying a Method Group”
on page 46, “Changing the Active Method Group” on page 47, or “Deleting a Method
Group” on page 47.
Parameter
Definition
Enable method group
filtering for users
Select to be able to create and define method groups.
Name
Choose a name for the method group.
Description
Provide a definition for the method group.
Available
Select methods to include in the group.
Selected
Show selected methods for the displayed group.
Analysis View
Use the Analysis view to view the reports, to process methods using sample results files, and to
view the data.
• Data Review Page
• Reports Page
• Real Time Status View
Data Review Page
Use the Data Review page to see initial information about a processed test. For more
information about using this page, see “Viewing Data Results” on page 60.
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Parameters
Description
File name
View the name of the data file.
Sample ID
View the ID of the sample.
Peaks
View peaks found in processing.
Retention Time
Specify the time after injection when an analyte elutes. This is the
total time that the analyte is retained on the chromatographic
column. If the maximum signal from an analyte is detected 5
minutes and 14 seconds after injection, then the analyte has a
retention time of 5:14.
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Analysis View
Parameters
Description
Relative Intensity (%)
View the intensity of the spectral peak used for observed molecular
weight value in target matching relative to the base peak in the
spectrum.
Data source
Choose a data source to process the data. See details on the “Peak
Processing Page” on page 104.
Spectrum
View the spectrum for the selected peak.
Elemental
Composition
View the results for elemental composition. Pathfinder checks the
most likely compound.
Target Matching/
Target Screening
View the results for target matching or target screening by clicking
the related tab.
Elemental Composition Values
Parameters
Description
Index
View the number for each formula candidate entry. The closest
entry has a check mark next to the index.
Mass
View calculated mass.
Formula
View the calculated formula for the compound.
RDB
View the ring and double bond (RDB) value to see which adducts
more closely match the value set.
Delta
View the distance of the formula candidate entry from the selected
elemental composition value.
Target Matching Values
Thermo Scientific
Parameters
Description
Apex RT
Retention time for the peak apex in minutes.
Height
Height of the peak.
Area
Area of the peak.
Observed molecular
weight
Value of the observed molecular weight.
Theoretical MW
Value of the theoretical molecular weight.
Difference (ppm)
Difference between the specified match and the identified peak.
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Analysis View
Reports Page
Use the Reports page to see report information about a processed test. For more information
about using this page, see “Viewing Reports” on page 67.
Parameters
Description
Report
Select the report name using the dropdown arrow.
Sample file
Select the name of the raw file that contains the sample data using
the dropdown arrow.
Real Time Status View
• Status View
• Sequence View
Status View
The status view displays the progress of the running sample. For information about using this
view, see “Viewing Instrument Status During Processing” on page 56. Details in this view can
change to support the instrument. Refer to the documentation that came with your
instrument for more information. Possible values include the following:
Parameter
Description
Status
View the current status of the instrument.
Scan speed
View the scan speed.
First scan
View the time for the first scan.
Last scan
View the time for the last scan.
Scan Number
View the number of the current scan.
Start time (min)
View the test start time in minutes.
Real time elapsed (min)
View the elapsed time from the start of the test in minutes.
Sequence View
The sequence view displays a list of sequences in the queue and their status. For information
about using this view, see “Viewing Instrument Status During Processing” on page 56.
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User Administration View
User Administration View
User View Page
User Mode Page
User View Page
For information about using these pages, see “Adding Users” on page 80, “Changing User
Information” on page 81, or “Configuring Instruments” on page 89.
Thermo Scientific
Parameter
Definition
Available Users area
View users that can log onto the User view of Pathfinder.
User Name
[Required] Type a single word (no spaces) identifying the
user.
Password
Type a password for the user.
Full Name
Type the first and last name of the user.
Phone number
Type the extension or phone number of the user.
Account number
Type the account number for the user.
Email address
[Required] Type the e-mail address of the user.
Notification allowed
Specify the type of notification the user can receive:
• None: The user receives no notification.
• E-mail only: The user receives e-mail notification only.
• E-mail and report: The user receives e-mail and reports
about test results.
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User Administration View
User Mode Page
To test an experiment, select the appropriate type of experiment in the User Mode area. For
information about using these pages, see “Working with Samples” on page 49.
• Survey Screening Page
• Target Matching Page
• Target Screening Page
Survey Screening Page
Parameter
Definition
User Name
Identify the person running the experiment.
Sample ID
Identify the sample.
Vial Position
Specify the position for the sample. If the vial position value
is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Submit. For more
information about the autosampler vial position format, see
your autosampler documentation.
File Name
Specify the file containing data for the sample.
Table
Sample ID
Identify the person running the experiment.
Vial
Specify the position for the sample. If the vial position value
is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Submit. For more
information about the autosampler vial position format, see
your autosampler documentation.
File Name
Specify the file containing data for the sample.
Method
Specify the method used for the experiment.
Method list area
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Method
Choose a method used for the experiment.
Method Description
If available, view a description for the method.
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User Administration View
Target Matching Page
Parameter
Definition
User Name
Identify the person running the experiment.
Sample ID
Identify the sample.
Vial Position
Specify the position for the sample. If the vial position value
is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Submit. For more
information about the autosampler vial position format, see
your autosampler documentation.
File Name
Specify the file containing data for the sample.
Target MW
Specify the molecular weight to look for in the sample.
Table
Sample ID
Identify the person running the experiment.
Vial
Specify the position for the sample. If the vial position value
is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Submit. For more
information about the autosampler vial position format, see
your autosampler documentation.
File Name
Specify the file containing data for the sample.
Target MW
Specify the molecular weight to look for in the sample.
Method list area
Thermo Scientific
Method
Choose a method used for the experiment.
Method Description
If available, view a description for the method.
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User Administration View
Target Screening Page
Parameter
Definition
User Name
Identify the person running the experiment.
Sample ID
Identify the sample.
Vial Position
Specify the position for the sample. If the vial position value
is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Submit. For more
information about the autosampler vial position format, see
your autosampler documentation.
File Name
Specify the file containing data for the sample.
Table
Sample ID
Identify the person running the experiment.
Vial
Specify the position for the sample. If the vial position value
is not in the correct format for your autosampler, Pathfinder
displays an error message when you click Submit. For more
information about the autosampler vial position format, see
your autosampler documentation.
File Name
Specify the file containing data for the sample.
Method
Specify the method used for the experiment.
Method list area
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Method
Choose a method used for the experiment.
Method Description
If available, view a description for the method.
Thermo Scientific
I
Index
A
D
A/D card data 25
abundance
intensity error weighted 35
mass error weighted 35
account name 2
active method group, changing 47
Active report 37
adding
hardware devices 90
peak 74
users 80
adducts 42
analog peak detection 105
analysis page 118
automatic file naming 94
data review
survey screening 61
target matching 62
Data review page 118
defining elemental composition values 33
deleting a user 81
Display options
Display 93
File Naming 94
Notification 97
System 96
double bonds 33, 110, 110
C
changing a method 43
changing test results
add a peak 74
manually integrating a peak 75
remove a peak 74
changing user information 81
chromatogram smoothing
Genesis 31
ICIS 30
commands 99
configuration files, settings 43
configure instruments 89
Create PDF 37
Create XML 37
creating a method 21
Thermo Scientific
E
elemental composition
display in data review 66
reference 109
values 32
Elemental Composition page 32, 66
entering user information 80
Expected RT 42
F
File menu 100
file names, automatic 94
fragments 42
G
General dialog box 103
General page, Method view 22
Genesis peak detection settings 108
Genesis Peak Integration 31
Genesis peak integration values 31
Go menu 101
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Index: H
H
O
hardware devices, adding 90
Help menu 101
High (ICIS), specifying algorithm 28
Open sequence dialog box, using 60
opening test results 60
options 99
Out Of Date Instrument Drivers Detected dialog box 117
I
ICIS peak detection settings 107
ICIS peak integration 30
instruments
establishing vial positions 92
instruments, configuring 89
M
manually integrating peaks 75
menus
File menu 100
Go menu 101
Help menu 101
Tools menu 100
method
changing 43
reviewing 43
testing 49
Method development option 102
method groups
activating 44
changing the active group 47
creating 44
deactivating 46
deleting 47
introduction 44
modifying 46
method type
survey screening 11
target matching 11
target screening 11
Method view, reference 102
Method view, using 22
methods, saving 37
MS processing, specifying values 33
multiple samples, templates for processing 54
N
new instrument, setting up tray and vial positions 92
Nitrogen Rule 33, 109
noise factors, ICIS 30
Notification view
changing 97
parameter values 97
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P
parameters, compound-specific 42
Pathfinder commands 99
Pathfinder options 99
PDA data 25, 104
peak detector algorithm, defining 28
peak parameters, defining 24
peak processing 104
peak tail integration
Genesis 31
ICIS 30
peak threshold, specifying 29
peaks
adding 74
manually integrating 75
removing 75
problems, troubleshooting 98
processing configuration file, setting parameters for 43
Processing page, Method view 24
R
remove a peak 75
reports
create PDF 37
create XML 37
define 37
print 37
Survey Screening 70
Target Matching 72
Target Screening long report 68
target screening summary report 67
Reports page
experiment results 70
reference 120
reprocessing
group of samples 77
single sample 76
results
elemental composition report 66
target matching 61
Retention Time Range, specifying 29
reviewing a method 43
ring equivalents 33, 110
Thermo Scientific
Index: S
S
U
sample, reprocessing 76
saving methods 37
sequence, reprocessing 77
setting up a virtual e-mail server 85
SMTP virtual e-mail server 85
Standard (Genesis), specifying algorithm 28
survey screening
data results 61
report 70
user process 11
Survey Screening page 50
System view, setting options 96
user
add new 80
changing information 81
deleting 81
entering information 80
user accounts, accessing Pathfinder 2
User view 80
UV channel 25
T
target matching
data results 62
report 72
results 61
user process 11
Target Matching page 50
target screening 38
long report 68
summary report 67
user process 11
Target Screening page 51
Target Screening report, view 67
Target Screening template, configuration file 41
templates for processing multiple samples 54
test report, view 70
testing a method 49
Tools menu 100
troubleshooting 98
Thermo Scientific
V
valley detection method 31
vial position, setting up for new instruments 92
view Target Screening report 67
view test report 70
view, setting display options 93
virtual e-mail server, setting up 85
W
workflow
Survey Screening 23
Target Matching 23
Target Screening 38
workflow results, viewing 59
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