Thermo Fisher Scientific BioPharma Finder 3.2 User guide

Thermo

BioPharma Finder

User Guide

Software Version 3.2

XCALI-98114 Revision A June 2019

BioPharma Finder, Exactive Plus, and Pinpoint are trademarks, and Chromeleon, Exactive, LCQ, LTQ, LTQ

FT, Orbitrap, Orbitrap Fusion, PepFinder, Thermo Scientific, and Xcalibur are registered trademarks of

Thermo Fisher Scientific Inc. in the United States.

The following are registered trademarks in the United States and other countries: Excel, Internet Explorer,

Microsoft, PowerPoint, and Windows are registered trademarks of Microsoft Corporation. Acrobat, Adobe,

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trademarks of Intel Corporation.

The following are registered trademark in the United States and possibly other countries: Mascot is a registered

service mark of Matrix Science Ltd. SQLite is a registered trademark of Hipp, Wyrick & Company, Inc.

ReSpect is a trademark of Positive Probability Ltd.

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.

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document shall in no way govern or modify any Terms and Conditions of Sale, which Terms and Conditions of

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Release history: Revision A, June 2019

Software version: Thermo BioPharma Finder 3.2 and later, Microsoft Windows 7 SP1 (64-bit), Microsoft

Windows 10 Professional (English) (64-bit)

For Research Use Only. Not for use in diagnostic procedures.

Thermo Scientific BioPharma Finder User Guide iii

C

Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xvii

Accessing Documentation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .xviii

Viewing the Product Manual. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .xviii

Viewing the Help and Animations. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .xix

Help Menu Commands. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .xix

System Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xx

Activating and Deactivating a License. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xx

Special Notices . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .xxi

Contacting Us . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xxii

Chapter 1 Introduction to BioPharma Finder . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1

BioPharma Finder Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

Protein Sequence Manager . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

Types of Analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

Using Common Features for All Types of Analyses . . . . . . . . . . . . . . . . . . . . . . . 4

Setting Up an Experiment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

Method Editor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

Run Queue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

Real-Time Optimization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

Starting the BioPharma Finder Application. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

Handling Database Service Error. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

Handling Other Errors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

Specifying Global Setting for Peptide Mapping Analysis . . . . . . . . . . . . . . . . . . 10

Specifying Global Settings for Intact Protein Analysis or Top Down

Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

Specifying the Default Raw Data File Folder . . . . . . . . . . . . . . . . . . . . . . . . . 14

Specifying the Image Dimensions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

Specifying the Precision for Mass Values . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17

Interacting with the User Interface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17

Exiting the BioPharma Finder Application . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

Backing Up Database and Files . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19

Retrieving Database and Files . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19

Managing Application Backup Using the Snapshot . . . . . . . . . . . . . . . . . . . . 20

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Data Conversion from Legacy Applications. . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

Converted Protein Sequences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22

Converted Processing Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Chapter 2 Using the Protein Sequence Manager and Editor. . . . . . . . . . . . . . . . . . . . . . . . . .25

Creating and Editing Protein Sequences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25

Importing a New FASTA File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26

Importing a New Sequence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29

Manually Creating a New Sequence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30

Editing a New Sequence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31

Editing the Amino Acids in an Existing Sequence . . . . . . . . . . . . . . . . . . . . . 33

Exporting a Sequence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34

Deleting an Existing Sequence. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35

Protein Sequence Manager Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . 35

Protein Sequence Editor Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37

Target Sequence Matching Components. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

Changing the Default Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40

Default Sublist of Modifications for Quick Loading . . . . . . . . . . . . . . . . . . . 40

Changing the Default and Visible Sublist of Modifications . . . . . . . . . . . . . . 40

Modification Assignments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

Managing Custom Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

Creating Custom Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

Modifying Custom Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46

Deleting Custom Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46

Modification Editor Pane Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

Saving a Protein Sequence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

Saving Sequence with the Same Name. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49

Saving Sequence with a Different Name . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49

Chapter 3 Assigning Modifications to a Protein Sequence . . . . . . . . . . . . . . . . . . . . . . . . . .51

Order of Modification Assignments. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52

Managing Disulfide Links . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52

Assigning Disulfide Links . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53

Removing Disulfide Links . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54

Disulfide Link Definitions Pane Parameters . . . . . . . . . . . . . . . . . . . . . . . . . 55

Managing Static Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55

Assigning Static Modifications. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55

Removing Static Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58

Residue Properties and Modifications Dialog Box Parameters . . . . . . . . . . . . 59

Managing Glycosylations. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60

Managing Variable Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62

Assigning Variable Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62

Variable Modifications for Intact and Peptide Analysis Pane Parameters . . . . 64

Contents

Thermo Scientific BioPharma Finder User Guide v

Managing Proteoforms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66

Defining the Modification List for Proteoforms . . . . . . . . . . . . . . . . . . . . . . 67

Generating and Saving the Proteoforms. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70

All Possible Proteoforms Table . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71

Chapter 4 Managing Theoretical Proteins and Peptides. . . . . . . . . . . . . . . . . . . . . . . . . . . . .73

Creating or Importing a Protein or Peptide Sequence . . . . . . . . . . . . . . . . . . . . 74

Choosing Digestion Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 76

Editing Target m/z Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78

Adding and Editing Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80

Managing the Processed Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83

Viewing the Processed Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84

Modifying the Results Display. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Saving the Processed Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Opening Previously Saved Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 86

Exporting the Processed Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87

Saving the Processed Results to a Workbook . . . . . . . . . . . . . . . . . . . . . . . . . 87

Amino Acid Letter Codes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90

Results Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 90

Chapter 5 Peptide Mapping Analysis Features. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .93

Experiment Results Display. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94

Quantification of Modifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94

Sequence Variant Analysis with Error-Tolerant Search . . . . . . . . . . . . . . . . . . . 94

De Novo Sequencing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95

Disulfide Mapping . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95

Localization of Glycosylation Sites on Glycopeptides . . . . . . . . . . . . . . . . . . . . 95

Running a Hydrogen Deuterium Exchange Experiment . . . . . . . . . . . . . . . . . . 96

HDX Deuterium Labeling. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97

Collecting HDX Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97

Processing an HDX Experiment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 98

HDX Output . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99

Glycan Structures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 100

Fragmentation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 103

Peptide Mapping Analysis Input . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 103

Peptide Mapping Analysis Output. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 104

Performing a Non-Targeted Peptide Mapping Analysis Experiment . . . . . . . . 104

Performing a Targeted Peptide Mapping Analysis Experiment . . . . . . . . . . . . 106

Chapter 6 Intact Protein Analysis Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .107

Deconvolution Algorithms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 108

Xtract Algorithm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109

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ReSpect Algorithm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109

Spectra Deconvolution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 110

Important Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111

Default Native Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111

Default Ion Trap Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112

Protein Quality Score . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112

Manual and Automatic Modes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112

Sliding Windows Deconvolution. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 113

Sliding Windows Advantages. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 114

Sliding Windows Steps . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115

Chromatographic Peak Detection and Spectral Peak Modeling. . . . . . . . . . . . 116

Batch and Multiconsensus Result Formats . . . . . . . . . . . . . . . . . . . . . . . . . . . 117

Target Sequence Matching . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 117

Extracted Ion Chromatogram Calculation for Deconvoluted Spectra . . . . . . . 119

Component XICs and Abundance Traces . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119

Drug-to-Antibody Ratio (DAR) Values. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120

Spectra Comparison . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120

Intact Protein Analysis Inputs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121

Intact Protein Analysis Outputs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121

Performing an Intact Protein Analysis Experiment . . . . . . . . . . . . . . . . . . . . . 121

Chapter 7 Top Down Analysis Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .125

Features Similar to Intact Protein Analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . 125

Top Down Analysis Inputs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126

Top Down Analysis Outputs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126

Performing a Top Down Analysis Experiment. . . . . . . . . . . . . . . . . . . . . . . . . 127

Chapter 8 Common Features for Different Analyses. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .129

Creating a New Experiment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129

Raw Data Files and Protein Sequences. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130

Loading the Raw Data Files. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130

Changing the Location of a Raw Data File . . . . . . . . . . . . . . . . . . . . . . . . . 133

Deleting the Raw Data Files . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134

Selecting One or More Protein Sequences . . . . . . . . . . . . . . . . . . . . . . . . . . 134

Importing a Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 136

Selecting a Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 137

Exporting a Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138

Deleting a Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139

Saving a Processing Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139

Navigating to the Method Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140

Method Summary Display. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140

Exporting the Method Summary and Saving the Method . . . . . . . . . . . . . . 143

Effects After Saving the Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144

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Using a Chromeleon-Compatible Workbook . . . . . . . . . . . . . . . . . . . . . . . . . 145

Managing a Workbook . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 145

Workbook Manager Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147

Importing and Exporting a Workbook . . . . . . . . . . . . . . . . . . . . . . . . . . . . 148

Editing a Workbook . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149

Workbook Editor Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 152

Chapter 9 Using the Run Queue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .159

Managing the Run Queue for Peptide Mapping Analysis . . . . . . . . . . . . . . . . 159

Pausing the Run Queue. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161

Resuming the Paused Queue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 162

Managing the Run Queue for Intact Protein Analysis . . . . . . . . . . . . . . . . . . . 162

Pausing the Run Queue. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 164

Resuming the Paused Queue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 165

Managing the Run Queue for Top Down Analysis . . . . . . . . . . . . . . . . . . . . . 165

Pausing the Run Queue. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167

Resuming the Paused Queue . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168

Using Common Run Queue Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168

Removing Selected Jobs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169

Removing Completed Jobs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169

Removing All Jobs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169

Queue Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 170

Chapter 10 Running a Peptide Mapping Analysis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .173

Data Acquisition and Peptide Identification . . . . . . . . . . . . . . . . . . . . . . . . . . 173

Starting a New Peptide Mapping Experiment . . . . . . . . . . . . . . . . . . . . . . . . . 174

Peptide Mapping Experiment Processing on the Queue Page . . . . . . . . . . . . . 177

Chapter 11 Working with a Peptide Mapping Processing Method . . . . . . . . . . . . . . . . . . . .179

Using a Processing Method for Peptide Mapping Analysis . . . . . . . . . . . . . . . 179

Editing Component Detection Parameters for Peptide Mapping Analysis . . . . 180

Editing the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 181

Component Detection Page Layout. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 181

Component Detection Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . 184

Viewing the Signal Threshold . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 188

Editing Identification Parameters for Peptide Mapping Analysis . . . . . . . . . . . 191

Editing the Identification Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192

Identification Page Layout. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192

Identification Page Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194

Editing HDX Parameters for Peptide Mapping Analysis . . . . . . . . . . . . . . . . . 201

Editing the Hydrogen Deuterium Exchange Page . . . . . . . . . . . . . . . . . . . . 201

Hydrogen Deuterium Exchange Page Layout . . . . . . . . . . . . . . . . . . . . . . . 202

Hydrogen Deuterium Exchange Page Parameters . . . . . . . . . . . . . . . . . . . . 204

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viii BioPharma Finder User Guide Thermo Scientific

Chapter 12 Viewing the Peptide Mapping Analysis Results. . . . . . . . . . . . . . . . . . . . . . . . . .207

Opening the Results from the Queue Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 207

Opening the Results from the Load Results Page. . . . . . . . . . . . . . . . . . . . . . . 209

Using Real-Time Optimization for Peptide Mapping Analysis . . . . . . . . . . . . 212

Viewing the Hydrogen Deuterium Exchange Plot. . . . . . . . . . . . . . . . . . . . . . 216

Opening the Hydrogen Deuterium Exchange Page . . . . . . . . . . . . . . . . . . . 216

Hydrogen Deuterium Exchange Page Display . . . . . . . . . . . . . . . . . . . . . . . 216

Hydrogen Deuterium Exchange Page Commands. . . . . . . . . . . . . . . . . . . . 217

Performing the Kinetic MS/MS Model Prediction . . . . . . . . . . . . . . . . . . . . . 218

Identifying Components Using De Novo Sequencing . . . . . . . . . . . . . . . . . . . 220

Performing De Novo Sequencing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 221

Canceling De Novo Sequencing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222

Setting Up the De Novo Sequencing Parameters. . . . . . . . . . . . . . . . . . . . . 222

Defining the Amino Acids for De Novo Sequencing . . . . . . . . . . . . . . . . . . 224

Organizing the Experimental Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 225

Chapter 13 Viewing the Process and Review Page for Peptide Mapping Analysis. . . . . .229

Process and Review Page Parameters for Peptide Mapping Analysis. . . . . . . . . 230

Process and Review Page Commands for Peptide Mapping Analysis . . . . . . . . 233

Viewing the Results Table for Peptide Mapping Analysis . . . . . . . . . . . . . . . . 234

Viewing the Peptide Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 235

Changing the Reference Condition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 235

Exporting the Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 236

Saving a Peptide Workbook from the Process and Review Page. . . . . . . . . . 237

Results Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 239

Modification Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 251

Results Table Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 252

Viewing the Chromatograms for Peptide Mapping Analysis . . . . . . . . . . . . . . 253

Viewing the Chromatograms. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 253

Chromatogram Plot Types . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 254

Displaying Multiple Chromatogram Plot Types for One File . . . . . . . . . . . 255

Displaying Same Chromatogram Plot Type for Multiple Files. . . . . . . . . . . 257

Chromatogram Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 259

Viewing the Trend Ratio Plot for Peptide Mapping Analysis. . . . . . . . . . . . . . 259

Viewing the Trend MS Area Plot for Peptide Mapping Analysis . . . . . . . . . . . 261

Viewing the Fragment Coverage Map for Peptide Mapping Analysis. . . . . . . . 262

Viewing the Fragment Coverage Map . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 263

Fragment Coverage Map Display . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 263

Viewing the Protein Sequence for Peptide Mapping Analysis . . . . . . . . . . . . . 265

Viewing the Protein Sequence . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 265

Protein Sequence Display . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 266

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Thermo Scientific BioPharma Finder User Guide ix

Viewing the Deconvoluted and Full-Scan MS Spectra for Peptide

Mapping Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 266

Viewing the Deconvoluted and Full-Scan MS Spectra. . . . . . . . . . . . . . . . . 267

Deconvoluted and Full-Scan MS Spectra Display . . . . . . . . . . . . . . . . . . . . 268

Full Scan Spectra Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 268

Viewing the Predicted and Experimental MS2 Spectra for Peptide

Mapping Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 268

Viewing the Predicted and Experimental MS2 Spectra . . . . . . . . . . . . . . . . 270

Predicted and Experimental MS2 Spectra Display . . . . . . . . . . . . . . . . . . . . 271

Predicted and Experimental MS2 Spectra Fragment Ions . . . . . . . . . . . . . . 272

MS2 Spectra Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 274

Comparing MS2 Spectra on the Process and Review Page . . . . . . . . . . . . . . 274

Switching the Header on the Process and Review Page . . . . . . . . . . . . . . . . 276

Manual Integration of Change Start and Stop Time of a Component . . . . . 277

Chapter 14 Viewing the Coverage Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .279

Coverage Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281

Viewing the Coverage Results Table . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281

Viewing the Results Table for Protein Coverage . . . . . . . . . . . . . . . . . . . . . 282

Exporting the Results Table Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 283

Results Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 283

Results Table Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 284

Viewing the Coverage Chromatogram. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 285

Viewing the Color-Coded Chromatogram. . . . . . . . . . . . . . . . . . . . . . . . . . 286

Modifying the Shading Settings. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 287

Chromatogram Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 288

Viewing the Coverage Map . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 289

Viewing the Sequence Coverage Map . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 290

Selecting the Sequence Coverage Map Components . . . . . . . . . . . . . . . . . . 290

Changing the Sequence Coverage Map Parameters . . . . . . . . . . . . . . . . . . . 291

Chapter 15 Viewing the Modification Summary Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .293

Modification Summary Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 295

Viewing the Modification Summary Results . . . . . . . . . . . . . . . . . . . . . . . . . . 297

Viewing the Modification Results Pane . . . . . . . . . . . . . . . . . . . . . . . . . . . . 299

Changing the Modification Summary Options . . . . . . . . . . . . . . . . . . . . . . 300

Exporting the Modification Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 302

Upper Table of Modification Results Pane Parameters . . . . . . . . . . . . . . . . 302

Lower Table of Modification Results Pane Parameters . . . . . . . . . . . . . . . . 303

Modification Results Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . 306

Contents

xBioPharma Finder User Guide Thermo Scientific

Viewing the Modification Summary Components. . . . . . . . . . . . . . . . . . . . . . 307

Viewing the Components Table . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 308

Changing the Abundance Calculation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 309

Exporting the Component Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 310

Saving a Peptide Workbook from the Modification Summary Page. . . . . . . 311

Components Table Commands. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 313

Viewing the Modification Plot . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 313

Comparing MS2 Spectra on the Modification Summary Page. . . . . . . . . . . 315

Switching the Header on the Modification Summary Page . . . . . . . . . . . . . 316

Chapter 16 Running an Intact Protein Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .319

Spectral Deconvolution for Intact Protein Analysis . . . . . . . . . . . . . . . . . . . . . 319

Starting a New Intact Protein Experiment. . . . . . . . . . . . . . . . . . . . . . . . . . . . 320

Selecting a Default Processing Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . 324

Differences Between Two Default Methods . . . . . . . . . . . . . . . . . . . . . . . . 325

Working in Manual Mode . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 326

Manual Mode Processing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 327

Deconvolving in Manual Mode. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 327

Chapter 17 Working with an Intact Protein Processing Method . . . . . . . . . . . . . . . . . . . . . .331

Using a Processing Method for Intact Protein Analysis . . . . . . . . . . . . . . . . . . 331

Editing Component Detection Parameters for Intact Protein Analysis. . . . . . . 332

Opening the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . . . 333

Left Side of the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . 333

Right Side of the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . 336

Editing the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 339

Chromatogram Parameters Area Parameters . . . . . . . . . . . . . . . . . . . . . . . . 342

Source Spectra Method Area Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . 345

Xtract Deconvolution Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 350

ReSpect Deconvolution Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 354

Component Detection Page Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . 363

Editing Identification Parameters for Intact Protein Analysis. . . . . . . . . . . . . . 363

Opening the Identification Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 364

Identification Page Layout. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 364

Editing the Identification Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 365

Left Side of the Identification Page Parameters . . . . . . . . . . . . . . . . . . . . . . 367

Right Side of the Identification Page Tables . . . . . . . . . . . . . . . . . . . . . . . . 369

Editing Report Parameters for Intact Protein Analysis . . . . . . . . . . . . . . . . . . . 370

Editing the Report Page. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 371

Report Page Layout . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 371

Report Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 372

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Thermo Scientific BioPharma Finder User Guide xi

Chapter 18 ReSpect and Sliding Windows Method Information . . . . . . . . . . . . . . . . . . . . . .375

Deconvoluted Spectra Display Mode. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 375

Optimizing the Protein Quality Score . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 377

Scoring Algorithm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 377

Specifying a Minimum Score. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 379

Viewing and Sorting the Scores . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 379

Model Mass Range Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 379

Best Results with the ReSpect Algorithm. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 380

Recommended Values for Sliding Windows Deconvolution Parameters . . . . . 381

Chapter 19 Viewing the Intact Protein Analysis Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . .383

Opening the Results from the Queue Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 383

Opening the Results from the Load Results Page. . . . . . . . . . . . . . . . . . . . . . . 384

Using Real-Time Optimization for Intact Protein Analysis . . . . . . . . . . . . . . . 388

Comparing Intact Protein Analysis Spectra . . . . . . . . . . . . . . . . . . . . . . . . . . . 392

Saving a Spectrum to the Library. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 393

Comparing Two Deconvoluted Spectra. . . . . . . . . . . . . . . . . . . . . . . . . . . . 394

Displaying Spectrum Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 395

Deleting Spectra from the Library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 396

Spectra Comparison Page Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 396

Spectra Comparison Page Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 397

Organizing the Experimental Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 398

Chapter 20 Viewing the Process and Review Page for Intact Protein Analysis . . . . . . . . .403

Process and Review Page Parameters for Intact Protein Analysis . . . . . . . . . . . 405

Process and Review Page Commands for Intact Protein Analysis. . . . . . . . . . . 407

Canceling Sliding Windows Processing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 408

Viewing the Results Table for Intact Protein Analysis . . . . . . . . . . . . . . . . . . . 409

Viewing the Intact Results Table . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 411

Exporting the Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 411

Saving an Intact Workbook. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 412

Results Table Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 414

Selecting a Reference Mass to Calculate Mass Differences . . . . . . . . . . . . . . 414

Viewing the Chromatograms for Intact Protein Analysis . . . . . . . . . . . . . . . . . 415

Viewing the Chromatograms. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 417

Chromatogram Pane Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 417

Chromatogram Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 418

Average Over RT Deconvolution and Auto Peak Detection

Chromatograms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 418

Sliding Windows Deconvolution Chromatograms . . . . . . . . . . . . . . . . . . . 419

Viewing the Deconvoluted Spectra for Intact Protein Analysis . . . . . . . . . . . . 420

Viewing the Deconvoluted Spectra . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 422

Deconvoluted Spectra Display for DAR . . . . . . . . . . . . . . . . . . . . . . . . . . . 423

Deconvoluted Spectrum Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . 424

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xii BioPharma Finder User Guide Thermo Scientific

Viewing the Source Spectra for Intact Protein Analysis . . . . . . . . . . . . . . . . . . 425

Viewing the Source Spectra . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 426

Source Spectrum Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 428

Viewing the Matched Sequence Information for Intact Protein Analysis . . . . . 428

Viewing the Matched Sequence Information. . . . . . . . . . . . . . . . . . . . . . . . 429

Component Information Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . 430

Target Match Sequence Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . 432

Viewing the Average DAR Values for Intact Protein Analysis . . . . . . . . . . . . . 432

Viewing the Average DAR Values . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 433

Average DAR Table Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 435

Component Specific Summary Table Parameters . . . . . . . . . . . . . . . . . . . . 435

Average DAR Pane Command . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 437

Chapter 21 Various Results Tables for Intact Protein Analysis . . . . . . . . . . . . . . . . . . . . . . .439

Results for Single-File/Batch Experiment Using Xtract and Average Over

RT Deconvolution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 439

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 440

Results Table Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 440

Results for a Single File/Batch Experiment Using Xtract and Auto Peak

Detection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 445

Results for a Single File/Batch Experiment using Xtract and Sliding

Windows Deconvolution. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 446

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 446

Results Table Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 447

Results for a Single File/Batch Experiment using ReSpect and Average

Over RT Deconvolution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 448

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 448

Results Table Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 450

Results for a Single File/Batch Experiment using ReSpect and Auto Peak

Detection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 451

Results for a Single File/Batch Experiment using ReSpect and Sliding

Windows Deconvolution. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 452

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 452

Results Table Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 453

Results for a Target Sequence Matching Experiment. . . . . . . . . . . . . . . . . . . . 454

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 455

Results Table Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 456

Results for a Multiconsensus Experiment . . . . . . . . . . . . . . . . . . . . . . . . . . . . 457

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 457

Results Table Columns . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 459

Results for a DAR-Enabled Experiment. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 464

Displayed Results Table. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 465

Results Table Additional Column . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 466

Contents

Thermo Scientific BioPharma Finder User Guide xiii

Chapter 22 Viewing an Intact Protein Analysis Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .467

Displaying a Report. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 468

Viewing Specific Sections in the Report. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 468

Saving a Report to PDF. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 469

Printing a Report. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 469

Reporting Page Toolbar. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 470

Report Sections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 470

Sample Information Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 471

Chromatogram Parameters Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 472

Chromatogram Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 473

Main Parameters Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 474

Advanced Parameters Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 475

Source Spectra Parameters Section. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 476

Sequences Information Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 476

Source Spectrum Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 477

Deconvoluted Spectrum Section . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 478

Masses Table Section. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 479

Component Detail Tables Section. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 481

Source Spectrum Evidence Plot Section. . . . . . . . . . . . . . . . . . . . . . . . . . . . 482

Chapter 23 Running a Top Down Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .485

Spectral Deconvolution for Top Down Analysis . . . . . . . . . . . . . . . . . . . . . . . 485

Starting a New Top Down Experiment. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 485

Top Down Experiment Processing on the Queue Page . . . . . . . . . . . . . . . . . . 488

Chapter 24 Working with a Top Down Processing Method . . . . . . . . . . . . . . . . . . . . . . . . . .489

Using a Processing Method for Top Down Analysis . . . . . . . . . . . . . . . . . . . . 489

Editing Component Detection Parameters for Top Down Analysis. . . . . . . . . 490

Opening the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . . . 491

Left Side of the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . 492

Right Side of the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . 494

Editing the Component Detection Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 497

Peak Selection Area Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 500

Fragmentation Types. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 504

Xtract Deconvolution Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 505

ReSpect Deconvolution Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 506

Component Detection Page Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . 506

Editing Identification Parameters for Top Down Analysis. . . . . . . . . . . . . . . . 507

Opening the Identification Page . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 508

Identification Page Layout. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 508

Editing the Identification Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 509

Left Side of the Identification Page Parameters . . . . . . . . . . . . . . . . . . . . . . 511

Right Side of the Identification Page Tables . . . . . . . . . . . . . . . . . . . . . . . . 511

Contents

xiv BioPharma Finder User Guide Thermo Scientific

Chapter 25 Viewing the Top Down Analysis Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .513

Opening the Results from the Queue Page . . . . . . . . . . . . . . . . . . . . . . . . . . . 513

Opening the Results from the Load Results Page. . . . . . . . . . . . . . . . . . . . . . . 514

Using Real-Time Optimization for Top Down Analysis . . . . . . . . . . . . . . . . . 517

Organizing the Experimental Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 520

Chapter 26 Viewing the Process and Review Page for Top Down Analysis . . . . . . . . . . . .525

Process and Review Page Parameters for Top Down Analysis . . . . . . . . . . . . . 527

Process and Review Page Command for Top Down Analysis . . . . . . . . . . . . . 529

Viewing the Results Tables for Top Down Analysis. . . . . . . . . . . . . . . . . . . . . 529

Viewing the Intact Fragmentation Results Table . . . . . . . . . . . . . . . . . . . . . 530

Exporting the Intact Fragmentation Results Table. . . . . . . . . . . . . . . . . . . . 531

Intact Fragmentation Results Table Parameters . . . . . . . . . . . . . . . . . . . . . . 531

Intact Fragmentation Results Table Commands . . . . . . . . . . . . . . . . . . . . . 535

Viewing the Intact Deconvolution Results Table. . . . . . . . . . . . . . . . . . . . . 536

Exporting the Intact Deconvolution Results Table . . . . . . . . . . . . . . . . . . . 537

Intact Deconvolution Results Table Parameters. . . . . . . . . . . . . . . . . . . . . . 538

Intact Deconvolution Results Table Commands . . . . . . . . . . . . . . . . . . . . . 539

Viewing the Chromatograms for Top Down Analysis . . . . . . . . . . . . . . . . . . . 540

Viewing the Chromatograms. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 541

Chromatogram Pane Options . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 542

Chromatogram Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 542

Viewing the Deconvoluted Spectra for Top Down Analysis . . . . . . . . . . . . . . 543

Viewing the Deconvoluted Spectra . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 544

Deconvoluted Spectra Panes Commands. . . . . . . . . . . . . . . . . . . . . . . . . . . 546

Viewing the Source Spectra for Top Down Analysis . . . . . . . . . . . . . . . . . . . . 548

Viewing the Source Spectra . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 549

Source Spectra Panes Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 551

Viewing the ProSightBP Fragment Map for Top Down Analysis . . . . . . . . . . 551

Viewing the ProSightBP Fragment Map . . . . . . . . . . . . . . . . . . . . . . . . . . . 553

Matching Fragment Detail Table Parameters. . . . . . . . . . . . . . . . . . . . . . . . 555

ProSightBP Fragment Map Pane Commands . . . . . . . . . . . . . . . . . . . . . . . 555

Viewing the ProSightBP Output Results for Top Down Analysis . . . . . . . . . . 556

Viewing the ProSightBP Output Results . . . . . . . . . . . . . . . . . . . . . . . . . . . 557

Exporting the ProSightBP Output Results. . . . . . . . . . . . . . . . . . . . . . . . . . 558

ProSightBP Output Pane Parameters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 558

ProSightBP Output Pane Commands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 562

Viewing the Matched Sequence Information for Top Down Analysis . . . . . . . 562

Modifying the Matched Sequence Information . . . . . . . . . . . . . . . . . . . . . . 563

Component Information Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . 564

Target Match Sequence Table Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . 564

Contents

Thermo Scientific BioPharma Finder User Guide xv

Appendix A Interactive Functions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .565

Rearranging the Panes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 565

Repositioning the Panes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 566

Rearranging the Panes with the Mouse . . . . . . . . . . . . . . . . . . . . . . . . . . . . 568

Collapsing the Panes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 569

Resizing the Panes Vertically . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 569

Resizing the Panes Horizontally. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 570

Using Basic Chromatogram Functions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 570

Zooming In. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 571

Zooming Out . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 572

Resetting to Original Scale . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 572

Copying to the Clipboard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 572

Displaying Labels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 573

Using Basic Spectrum Functions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 573

Zooming In. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 573

Zooming Out . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 575

Resetting to Original Scale . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 575

Copying Spectrum to the Clipboard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 575

Copying Data to the Clipboard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 575

Using Copy and Paste Functions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 576

Copying to an Excel File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 577

Copying a Portion of the Pane. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 578

Using Basic Table Functions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 578

Sorting Rows . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 578

Showing or Hiding Columns. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 579

Filtering Data in a Table . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 580

Filtering Data Rows. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 581

Selecting Filter Operators . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 582

Table Filter Operators . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 583

Selecting Filter Operands. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 585

Table Filter Operands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 586

Setting Up Custom Filters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 586

Removing One Filter. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 587

Removing All Filters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 588

Saving Filters to a File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 588

Applying Saved Filters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 588

Appendix B Glycans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .589

N-Linked Glycans in the Define Modification List Window. . . . . . . . . . . . . . 590

N-Linked Glycans with a CHO Host Cell-Line Type . . . . . . . . . . . . . . . . . . . 597

N-Linked Glycans with a Human Host Cell-Line Type . . . . . . . . . . . . . . . . . 605

O-Linked Glycans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 607

Contents

xvi BioPharma Finder User Guide Thermo Scientific

Appendix C References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .609

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .611

Thermo Scientific BioPharma Finder User Guide xvii

P

Preface

This guide describes how to use the Thermo BioPharma Finder™ 3.2 application to

characterize biotherapeutic proteins by using three key workflows: Peptide Mapping Analysis,

Intact Protein Analysis, and Top Down Analysis.

To suggest changes to the documentation or to the Help

Complete a brief survey about this document by clicking the button below.

Thank you in advance for your help.

Contents

•Accessing Documentation

•System Requirements

•Activating and Deactivating a License

•Special Notices

•Contacting Us

Preface

xviii BioPharma Finder User Guide Thermo Scientific

Accessing Documentation

In addition to this guide, the BioPharma Finder application includes Help, animations, and

the Thermo BioPharma Finder Installation Instructions, available on the software DVD.

For more details about how to access the documentation, see the following topics:

•Viewing the Product Manual

•Viewing the Help and Animations

•Help Menu Commands

Viewing the Product Manual

You can view the product manual either from the BioPharma Finder application or from the

Thermo Fisher Scientific™ website.

Table 1. Viewing the product manual

To view from … Do this …

The application From the BioPharma Finder window, choose Help > Manual >

BioPharma Finder User Guide.

The website 1. Go to thermofisher.com.

2. Point to Services & Support and click Manuals on the left.

3. In the Refine Your Search box, search by the product name.

4. From the results list, click the title to open the document in

your web browser, save it, or print it.

To return to the document list, click the browser Back

button.

Preface

Thermo Scientific BioPharma Finder User Guide xix

Viewing the Help and Animations

You can view both the Help and animations from the BioPharma Finder application.

Help Menu Commands

The Help menu in the BioPharma Finder application provides the following commands.

Table 2. Viewing the Help and animations

To view … Do this …

The Help From the BioPharma Finder window, choose Help > BioPharma

Finder Help. To locate a particular topic, use the Help Contents,

Index, or Search panes.

In applications that have a Communicator bar, click the field or

parameter to display definitions, required actions, ranges, defaults, and

warnings, if they are available.

Note Messages in the Communicator bar sometimes do not

synchronize with the current field after you move to another user

interface area and then move back.

The animations From the BioPharma Finder window, choose Help > Animations.

Then, view a specific animation by clicking its corresponding link.

Help menu commands Description

Snapshot Creates and restores backup of the application files and

databases.

BioPharma Finder Help Displays the BioPharma Finder Help.

How to Use Help Displays the BioPharma Finder Help topic that explains

how to use the Help.

Glossary Displays pages of glossary terms in alphabetical order and

their related definitions.

Manual Provides access to the BioPharma Finder User guide in

PDF format.

Animations Displays links to animations that demonstrate typical user

interactions in various views.

About BioPharma Finder Displays the release and copyright information. Provides

access to the license activation/deactivation dialog box,

the version information, and the user license agreement

for the BioPharma Finder application.

Report an Issue Provides access to the submission of defects or

enhancement requests.

Preface

xx BioPharma Finder User Guide Thermo Scientific

System Requirements

The following are the minimum and recommended system requirements for BioPharma

Finder 3.2 operation.

Activating and Deactivating a License

Use the Thermo Scientific™ Product Licensing wizard to activate or deactivate the license for

the BioPharma Finder application. To activate the license, you must have an activation code

from Thermo Fisher Scientific. You must deactivate the license before you transfer it to

another computer.

To start the license activation or deactivation process

1. Open the BioPharma Finder application.

2. Choose Help > About BioPharma Finder to display the About dialog box.

3. Click Activate (Deactivate) to start the activation (or deactivation) process, as applicable.

4. Follow the instructions in the License Activation wizard.

For additional instructions, click Help in the wizard.

System Minimum requirements Recommended requirements

Hardware • Intel Core™ i7-4770 CPU@3.40 GHz

• 8 GB registered RAM

• 100 GB storage hard drive—

ST1000DM-003 SCSI disk device

•DVD/CD-ROM drives

• Resolution display 1280 1024

(SXGA)

• Quad-core Intel™ Xeon™ CPU (E5-1630 v3

3.7 GHz 10 MB 2133 4C)

• 32 GB DDR4-2133 (4 8 GB) registered RAM

• 2 TB storage hard drive (SATA, 7200 rpm)—

512 GB solid state boot drive (SATA)

• DVD-RW optical drive

• Resolution display 1920 1080 (WUXGA)

Software • Microsoft™ Windows™ 7 Professional

(English) SP1 (64-bit)

• Microsoft .NET Framework 4.6.2

• Microsoft Office 2010

• Adobe™ Acrobat Reader™ DC

• Microsoft Windows 10 Professional (English)

(64-bit)a

• Microsoft .NET Framework 4.6.2

• Microsoft Office 2016

• Adobe Acrobat™ Pro DC

aWindows 10 Enterprise LTSB edition is also supported.

Tip If you are not able to see the entire interface of the BioPharma Finder application,

make sure that your computer resolution is set to at least 1280 1024.

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Thermo Fisher Scientific BioPharma Finder 3.2 User guide

Thermo Fisher Scientific BioPharma Finder 3.2 User guide

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