Manuals - Thermo Fisher Scientific

Manuals - Thermo Fisher Scientific
User Manual
OncoScan Console 1.1
For Research Use Only. Not for use in diagnostic procedures.
P/N 703195 Rev 2
Affymetrix® OncoScan™ Console 1.1
Trademarks
Affymetrix®, OncoScan™ ,GeneChip®, NetAffx®, Command Console®, Powered by Affymetrix™, GeneChip-compatible™, Genotyping
Console™, DMET™, GeneTitan®, Axiom®, CytoScan®, and GeneAtlas® are trademarks or registered trademarks of Affymetrix, Inc. All other
trademarks are the property of their respective owners.
All other trademarks are the property of their respective owners.
This database/product contains information from the Online Mendelian Inheritance in Man® (OMIM®) database, which has been obtained
under a license from the Johns Hopkins University. This database/product does not represent the entire, unmodified OMIM® database,
which is available in its entirety at http://www.omim.org/
Limited License Notice
Limited License. Subject to the Affymetrix terms and conditions that govern your use of Affymetrix products, Affymetrix grants you a nonexclusive, non-transferable, non-sublicensable license to use this Affymetrix product only in accordance with the manual and written
instructions provided by Affymetrix. You understand and agree that except as expressly set forth in the Affymetrix terms and conditions, that
no right or license to any patent or other intellectual property owned or licensable by Affymetrix is conveyed or implied by this Affymetrix
product. In particular, no right or license is conveyed or implied to use this Affymetrix product in combination with a product not provided,
licensed or specifically recommended by Affymetrix for such use.
Patents
Software products may be covered by one or more of the following patents: U.S. Patent Nos. 5,733,729; 5,795,716; 5,974,164; 6,066,454;
6,090,555; 6,185,561; 6,188,783; 6,223,127; 6,228,593; 6,229,911; 6,242,180; 6,308,170; 6,361,937; 6,420,108; 6,484,183; 6,505,125; 6510,391;
6,532,462; 6,546,340; 6,687,692; 6,607,887; 7,062,092 and other U.S. or foreign patents.
Copyright
© 2014 Affymetrix, Inc. All rights reserved.
Contents
Chapter 1
3
Installation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
System Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
Installing OncoScan Console . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
Starting OncoScan Console . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .6
Chapter 2
First Time Setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Setting Up OncoScan Console . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Assigning Input Sample Paths . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Assigning New Input Sample Paths . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
Using Existing Input Sample Paths . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Assigning an Output Results Path . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Assigning a New Output Results Path . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Using an Existing Output Results Path . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
Adding Sub-Folders . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
Assigning a Central QC History Path . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
Assigning a Library File Path . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
Downloading Analysis Files from NetAffx . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
Setting Proxy Server Access . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .13
Un-installing OncoScan Console . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14
Chapter 3
Standard Analysis Setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
Setting Up a Standard Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
Confirming or Changing Array Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .16
Adding CEL Files to Analyze . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Manually Adding CEL Files to Analyze . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
CEL File Displaying Options (Optional) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Importing CEL Files Using Batch Import . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Generating Result File Names . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 22
Correcting Mismatched CEL File Pairings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Using the Sorting Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Generating a Result File Name after Sorting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Setting your Output Information Location (Optional) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Selecting a Suffix to Append to the Analysis Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24
Exporting Batch Analysis Files (Optional) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 25
Log File Collection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
Viewing the Log Collection File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
Log Rollover . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
Chapter 4
Analysis Submission and QC Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Submitting your Analysis Setup Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
Workflow Dashboard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29
QC Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Interpreting and Using QC Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .30
Customizing QC Metrics and Thresholds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30
Contents
4
Exporting the QC Results Table . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
Generating and Exporting Reports . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .34
CelPairCheck Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34
Gene Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37
Probe Level Data Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41
Segment Data Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 44
Somatic Mutation Data Report . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47
Export All Data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51
Chapter 5
Matched Normal Analysis Setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Setting Up a Matched Normal Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
Confirming or Changing Array Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .54
Adding CEL Files to Analyze . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
Manually Adding CEL Files to Analyze . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
CEL File Displaying Options (Optional) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57
Importing CEL Files Using Batch Import . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
Generating Result File Names . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
Correcting Mismatched CEL File Pairings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Using the Sorting Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Generating a Result File Name after Sorting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Setting your Output Information Location (Optional) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Selecting a Suffix to Append to the Analysis Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 62
Exporting Batch Analysis Files (Optional) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63
Log File Collection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 64
Viewing the Log Collection File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65
Log Rollover . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66
Appendix A
Appendix: Custom Reference Files . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
Creating your own Reference File . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
Appendix B
Appendix: QC Metrics - Definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68
Array Data QC Metrics (Overview) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68
MAPD (Median of the Absolute Values of all Pairwise Differences) . . . . . . . . . . . . . . . . . 68
ndSNPQC (SNP Quality Control of Normal Diploid Markers) . . . . . . . . . . . . . . . . . . . . . . . 68
SNP QC Type (SNP Quality Control Type) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68
CelPairCheck Status . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68
CelPairCheck Compare Rate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68
CelPairCheck Concordance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
ndWavinessSD (Normal Diploid Waviness Standard Deviation) . . . . . . . . . . . . . . . . . . . . .69
Y Gender Call . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .69
ndCount . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
Low Diploid Flag . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
ACDC (Aberrant Cell-Derived Copy Number) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
%Aberr. Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .70
TuScan Ploidy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Offset Flag . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Contents
5
L2R Offset . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Low % Aberrant Cell nGoF . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Hyb Control Intensity_AT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Hyb Control Intensity_GC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Q3 Raw Intensity_AT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 70
Q3 Raw Intensity_GC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
AGR_AT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
AGR_GC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
ndSNR_AT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
ndSNR_GC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .71
ndRawSNPQC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Call Rate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Matched Normal Compare Rate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 71
Matched Normal Concordance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72
Array Data QC Metrics (Detailed Descriptions) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .73
MAPD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .73
Effect of MAPD on Functional Performance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74
ndWaviness-SD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .74
ndSNPQC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 75
Effect of ndSNPQC on Functional Performance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 75
CelPairCheckStatus . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 77
ndWavinessSD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78
% Aberrant Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78
Low Diploid Flag . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 78
Appendix C
Appendix: Algorithms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79
B-allele Frequencies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79
LOH Algorithm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79
TuScan Algorithm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .80
OncoScan™ Console 1.1 User Manual
Installation
System Requirements
Operating System
Windows® 7 Professional (64-bit) with Service Pack 1 installed
Installing OncoScan Console
To install OncoScan Console:
1. Go to www.affymetrix.com and navigate to the following location:
Home > Products > Microarray Solutions > Instruments and Software >
Software >
2. Locate and download the zipped OncoScan Console software package.
3. Unzip the file, then double-click OncoScanSetup64.exe to install it.
4. Follow the directions provided by the installer.
Starting OncoScan Console
To start OncoScan Console:
1. Locate the OncoScan Console Shortcut
on your system’s Desktop, then double-
click on it.
A window appears prompting you to set your Library path. (Figure 1.1)
Figure 1.1 Set Library Path message
2. Click OK.
A File window appears. (Figure 1.2 on page 7)
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OncoScan™ Console 1.1 User Manual
7
Figure 1.2 File window
3. Navigate to your Library File folder location, then click OK or click Create New Folder
to create a new library folder (Example: OncoScan Analysis Library Files).
The OncoScan application opens. (Figure 1.3 on page 8)
OncoScan™ Console 1.1 User Manual
Figure 1.3 OncoScan - Main window
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OncoScan™ Console 1.1 User Manual
9
First Time Setup
Setting Up OncoScan Console
The first time you launch OncoScan Console, the following message appears: (Figure 2.1).
Figure 2.1 Set Paths message
Acknowledge the message, click OK, then do the following:
NOTE: You only need to perform the following section once, as the data and selections
you input (throughout this section) are retained for your convenience.
Assigning Input Sample Paths
The Input Sample Path folder is the location you normally store your CEL files.
Assigning New Input Sample Paths
To assign new input sample paths:
1. Click Add.
OncoScan™ Console 1.1 User Manual
The following window appears: (Figure 2.2)
Figure 2.2 Add Input sample files window
2. Navigate to the recommended C:\Users directory, then click the Create New folder.
3. In the Create New Folder field, enter a folder name.
(Example: C:\Users\YourName\OncoScan_CEL_files)
4. Click OK.
Your new folder appears.
Using Existing Input Sample Paths
To use existing input sample paths:
1. Click Add.
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OncoScan™ Console 1.1 User Manual
The following window appears: (Figure 2.2)
Figure 2.3 Add Input sample files window
2. Navigate to your desired input sample path location, then click OK.
Assigning an Output Results Path
Assigning a New Output Results Path
To assign a new Output Results Path:
1. Click the Output results path field’s browse button.
An Explorer window appears.
2. Navigate to the recommended C:\Users directory, then click Create New Folder.
3. In the Create New Folder field, enter a folder name.
(Example: C:\Users\YourName\OncoScan_results_files)
4. Click OK.
5. Click Save.
Using an Existing Output Results Path
To use an existing Output Results Path:
1. Click the Output results path field’s browse button.
An Explorer window appears.
2. Navigate to the desired output sample path location, then click OK.
3. Click Save.
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OncoScan™ Console 1.1 User Manual
12
Adding Sub-Folders
TIP: To better organize your output results, you can add sub-folders to your newly
assigned output result path’s folder.
To add sub-folders to your newly assigned result path’s folder:
1. The Output results path field’s browse button to return to your assigned output path
and/or folder.
2. Click Create New Folder.
3. Enter a sub-folder name.
4. Click OK.
The newly created sub-folders now appear in the output result information window.
5. Repeat the above steps 1-4 to add more sub-folders.
Assigning a Central QC History Path
To assign a Central QC history path:
1. Click the Central QC history path field’s browse button.
An Explorer window appears.
2. Navigate to: C:\ProgramData\Affymetrix\OncoScan
3. Click Create New Folder (lower left) to create a Central QC history path folder.
4. In the Create New Folder field, enter a folder name. (Example: My_QC_History)
5. Click OK.
6. Locate your new folder, single-click on it, then click OK.
Your new folder information now appears in the Central QC History path field.
Assigning a Library File Path
Make sure your assigned Library Path folder is placed in a high-level, easy to access, local
directory. (Example: C:\)
To assign a Library File path:
1. Click the Library File path field’s browse button.
An Explorer window appears.
2. Navigate to: a high-level, easy to access, local directory.
(Example: C:\)
3. Click Create New Folder (lower left) to create a Library Files path folder.
4. In the Create New Folder field, enter a folder name. (Example: C:\OncoScanLib)
5. Click OK.
6. Locate your new folder, single-click on it, then click OK.
Your new folder information now appears in the Library file path field.
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Downloading Analysis Files from NetAffx
After your Library Path folder is created, you must download the library files that OncoScan
Console uses to analyze and annotate the data from NetAffx.
NOTE: You can also download the analysis library file package from directly from
www.affymetrix.com. After downloading, unzip the contents of the file directly into the
Library folder you assigned earlier. See Assigning a Library File Path on page 12.
If you go to the website (outside of OncoScan Console) to download the analysis library
file package, you must close, then restart OncoScan Console in order for it to recognize
the newly downloaded files.
To download OncoScan analysis files from NetAffx:
1. Click on Utility Actions -> Download Library Files or open your Internet browser and
go to www.affymetrix.com.
2. Enter your NetAffx user name and password or click Register Now to create a NetAffx
account.
The Choose Files window opens with a list of array types supported by the software.
3. Click the OncoScan array checkbox.
4. Click Next.
The Download Progress window displays the progress of the downloading and
unpacking of the files.
Setting Proxy Server Access
This configuration should only be done if the user’s system has to pass through a proxy
server to access Affymetrix NetAffx server.
In most cases, when a customer requires the use of a proxy, they can set a system-level
proxy using their default Internet browser while keeping the OncoScan Console default
setting at Use System Proxy.
NOTE: Please contact your IT department if you do not know what proxy settings you need.
To set the Proxy Server Access in OncoScan Console:
1. Click the Utility Actions button (upper right).
2. From the Configuration window, click Set Proxy Server (lower right).
The Download Library Files window opens (Figure 2.4).
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Figure 2.4 Configuration window
3. Click the Proxy Server checkbox, then complete the required fields.
4. Click Save.
NOTE: This proxy user ID and password is NOT the same ID and password used to connect
to the Affymetrix NetAffx server.
Un-installing OncoScan Console
1. From the Windows Start Menu, navigate to the Windows Control Panel.
2. Navigate to the Uninstall or change a program.
3. Locate the OncoScan Console application, then perform the un-install as you normally
would.
TIP: You can also un-install OncoScan by re-running the OncoScanSetup64.exe installation
file.
NOTE: Your data and library files are NOT deleted by uninstalling OncoScan Console.
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Standard Analysis Setup
To setup a Matched Normal Analysis (See Matched Normal Analysis Setup, on page 53)
Setting Up a Standard Analysis
Figure 3.1 Analysis Setup window/tab - Overview
As long as your library file folder contains the necessary analysis files for the array, your
configuration paths are established and your Array Information fields auto-populate.
(Figure 3.2)
Figure 3.2 Standard Analysis Configuration
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Confirming or Changing Array Information
1. From the Select array type drop-down list, select OncoScan.
NOTE: The Select array type drop-down list includes only the array types from the library
(analysis) files that have been downloaded from NetAffx or copied from the Library
package provided in the OncoScan installation package.
IMPORTANT: After adding new library files to the library file folder, always close and relaunch OncoScan Console to ensure the newly added files are recognized by the software.
2. From the Select analysis workflow drop-down list, click to select an analysis workflow.
By default, the Analysis Workflow is set to FFPE Analysis NA33.
Other available Analysis Workflow options are:
Ref103 Analysis NA33 - Use this workflow with cell line DNA.

FFPE Analysis including Matched Normal NA33 - Use this workflow when

you have DNA from normal and tumor tissue from the same FFPE fixed specimen.
(See Setting Up a Matched Normal Analysis, on page 53)
Reference Generation NA33 - Select this option when you want to create your

own Reference File. See Appendix A: Appendix: Custom Reference Files, on
page 67.
3. Enter a Workflow name (optional). By default, the Set workflow name is Workflow.
Click
(upper right) to enter a different workflow name.
TIP: Customizing a Workflow name can be a useful tool in keeping track of analysis
workflows as all the related output files (outside of the OSCHP file) are pre-fixed with this
workflow name.
The Annotation file is automatically selected for you and is based on your selected
reference model file. (Example: OncoScan.na33.v1.annot.db)
NOTE: The Annotation to be used for analysis field is auto-populated based on your Ref
Model file selection. The analysis is not be permitted to run if the appropriate annotation
file is not available in your Library folder.
4. Select a Somatic mutation reference model file. By default, it is set to
OncoScan.na33.v1.SOM_REF_MODEL or the most recently used SOM reference
model file. If you created your own reference model file, click the drop-down list to
select your .SOM_REF_MODEL.
5. Confirm the displayed Somatic mutation threshold file to be used is correct. If you need
to change it, click the Browse button, navigate to the appropriate threshold .txt file,
then click OK.
IMPORTANT: If the Reference Model File and Somatic mutation Reference Model File
were created independently of each other, a warning message appears after you click
Submit (to start the Workflow Analysis process). Click OK to acknowledge the message.
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Adding CEL Files to Analyze
You can manually add CEL files or import them as a tab-delimited text file.
Manually Adding CEL Files to Analyze
To add a batch file containing the list of CEL files, see Importing CEL Files Using Batch
Import, on page 20.
To manually add CEL files:
1. From the Select the intensity (CEL) file(s) to analyze pane, click the Add CEL files drop-
down.
2. Click AT Channel.
The CEL file window appears. (Figure 3.3 on page 17).
NOTE: Immediately after a CEL file has been submitted for analysis (within the OncoScan
Console software), it goes into a locked state and cannot be used for other analyses until
the current analysis is complete.
Locked CEL file.
Figure 3.3 CEL file folder -EXAMPLE
IMPORTANT: Affymetrix recommends using an “A” or “C” as the last character to
designate the channel in the CEL file naming convention. Example: “_AS_05A.CEL” is an
AT Channel file, while “_AS_05C.CEL” is a GC Channel file. See Figure 3.3.
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3. Click any header to sort your files or click the Files of type drop-down to filter your CEL
files by AT Channel, as shown in Figure 3.4.
Figure 3.4 Files of type drop-down list
4. Single click, Ctrl click, or Shift click (to select multiple AT Channel files)
5. Click Open.
The AT Channel fields are now populated. (Figure 3.5)
Figure 3.5 AT Channel file list
6. Click the Add CEL files drop-down.
7. Click GC Channel. The CEL file window appears. (Figure 3.3)
8. Click any header to sort your files or click the Files of type drop-down to filter your CEL
files by GC Channel, as shown in Figure 3.6.
Figure 3.6 Files of type drop-down list
9. Single click, Ctrl click, or Shift click (to select multiple GC Channel files).
10. Click Open.
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The GC Channel fields are now populated. (Figure 3.7)
Figure 3.7 CEL Files Loaded to Analyze
CEL File Displaying Options (Optional)
The File Name drop-down list (Figure 3.8) is dynamically populated and based on what
attributes are populated in the ARR file.
To use this display option, you must:
1. Provide the appropriate attributes at the time of sample registration in AGCC.
2. The ARR files must reside in the same folder as the CEL files.
Figure 3.8 EXAMPLE: File Name
Display Choices
To see “channel” (as an option in the drop down), you must use a template (or the
OncoScan template provided in the library files) that contains a “channel” attribute. The
resulting ARR file must also reside in the same folder as the CEL files you are analyzing.
You can display one of the attributes from the ARR file in the table. For example,
“Channel” can be chosen (Figure 3.8) to confirm the assignment of a CEL file to its
appropriate channel.
To select a FIle Name display attribute:
1. Click the File Name drop-down button, then click to select the attribute you want
displayed along with your CEL file names.
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The two examples (Figure 3.9 and Figure 3.10) show how the table appears with the
display set to Filename, then to Channel.
Figure 3.9 Table with Filename displayed
Figure 3.10 Table with Channel displayed
Importing CEL Files Using Batch Import
OncoScan Console allows import of CEL files using a batch file. The batch file must be saved
as a text (Tab-delimited) format and include the full directory path for your CEL files (as
shown in Figure 3.11).
TIP: The resulting OSCHP files are saved to your output path location, therefore it is not
necessary to include a path under RESULT. Simply enter the desired results filename in this
column.
The format for this tab-delimited file is 3 columns (A,B, and C) with the headers:
ATCHANNELCEL

GCCHANNELCEL

RESULT

OncoScan™ Console 1.1 User Manual
You must provide the full path to the CEL files for each Channel column.
(Example: C:\Desktop\OncoScan\Data\Sample1.cel)
Figure 3.11 List from Microsoft Excel
1. Click Import Batch File
A File window appears.
2. Navigate to your text (tab-delimited) file location, then click on the file you want to
import.
IMPORTANT: The Microsoft Excel application must be closed before you import (click
Open).
3. Click Open.
The AT, GC, and Result File Name fields are now populated. (Figure 3.12)
Figure 3.12 Tab-delimited text file imported into OncoScan Console
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Generating Result File Names
Results File Names can either be entered in manually or OncoScan Console can generate
them automatically.
NOTE: If you use the suffix option (Selecting a Suffix to Append to the Analysis Results,
on page 24) and enter your Result File Names manually, your assigned suffix appears in
the Results File Name column.
If you auto-generate your Results File Names, your assigned suffix appears in the Results
File Name column, but it does get added to your final OSCHP file name(s).
To manually enter a Results File Name:
1. Single-click inside the appropriate Results Name File field to produce a cursor, then type
in the file name you want.
To auto-generate a suggested Result File Name:
NOTE: During the Result File Name auto-generation process, the file names are compared
to identify their common root name for use as a results file name. Generally, the last 5
characters of each CEL file name are ignored, then the remaining root names of the AT
and GC file names are compared. If the root names of the AT and GC channel match, then
the root name is used in the Results File Name field. The one exception is if your array
name “_(OncoScan)” is appended to the file name during registration in Affymetrix
GeneChip Command Console (AGCC). In this case, the “_(OncoScan)” is ignored during the
comparison, but then added back in the Results File Name field.
1. After the AT and GC Channel lists are populated, click the Result File Names drop-
down, then select Auto Generate Output Name.
2. The Result File Name column is now populated with suggested filenames for each
pairing. (Figure 3.13)
Figure 3.13 Result File Name list
Common root names should be consistent all the way up to the last character of the CEL
file name prior to the .cel extension. If there is a paired file mis-match, the Results File
Name appears as Output1. (Figure 3.14)
Figure 3.14 Result File Name “Output”
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If Output1 or subsequent Outputs (Output 2, Output 3...) appear, investigate the validity
of your original pairing. See Correcting Mismatched CEL File Pairings, on page 23.
To edit an auto-generated Result File Name:
1. Click on the Result File name you want to edit.
2. After the cursor appears, edit the filename as you normally would.
3. Click outside the row to save your edit.
To clear the entire Result File Name column:
1. Click the Result File Names drop-down button, then select Clear Column.
The column is now cleared and ready for new Result File Name entries.
Correcting Mismatched CEL File Pairings
If there is a paired file mismatch, the Results File Name appears as Output1, Output2,
Output3, etc.
A paired file mismatch is most likely caused by an incorrect CEL filename pairing and not a
mismatch of your native CEL files.
A simple way to correct mismatches is to sort the AT and GC columns so that files with the
same root names are next to each other.
TIP: Common root names should be consistent all the way up to the last character of the
cel file name prior to the .cel extension. Affymetrix recommends using an “A” or “C” as
the last character to designate the channel in the CEL file naming convention. Example:
“_AS_05A.CEL” is an AT Channel file, while “_AS_05C.CEL” is a GC Channel file.
Using the Sorting Features
To sort an individual column:
1. Click on either the AT or GC Channel header.
The column is now sorted in an ascending order.
2. Click on either the AT or GC Channel header again to reverse the sorting order.
To sort both columns simultaneously:
1. Click Sort All.
The contents of each column are now sorted together in an ascending order.
2. Click Sort All again.
The contents of each column are now sorted together in a descending order.
To swap CEL files between columns:
1. Click and drag a column CEL entry onto another column CEL entry, then release the
mouse button.
The CEL entries have now swapped column positions.
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To reorder the CEL files in a column:
1. Click and drag a CEL file to another position within the column, then release the mouse
button.
The CEL file is now at its new position.
To add a cell to a column:
1. Click and drag a column cell to the top or bottom border line of a neighboring cell, then
release the mouse button.
Generating a Result File Name after Sorting
1. After your AT and GC Channel lists are properly sorted, click the Result File Names
drop-down, then select Auto Generate Output Names.
The Result File Name column is now populated with suggested filenames for each
pairing.
If OncoScan Console detects an inconsistency between the AT and GC file names to be
paired, a Result File Name labeled, “Output n” reappears.
IMPORTANT: Confirm that both columns are sorted in the same direction. If they are,
examine the files and confirm they are paired correctly. The file names (excluding the last
character before the .CEL) MUST match exactly.
Repeat the sorting steps above, then try to Auto Generate Output Names again until a
successful Result File Name(s) appears.
Setting your Output Information Location (Optional)
The Output result information path (lower left) is retained from your initial setup.
To select a different folder to store your results:
1. Click the browse button, then navigate to the folder you want. If you want to change
the default folder, see Assigning an Output Results Path, on page 11.
Selecting a Suffix to Append to the Analysis Results
You can append a suffix at the end of all your Results File Names. This is useful when
tracking versions of the analysis files used to generate the resulting OSCHP files.
To use an appending suffix:
1. Click inside the Select a suffix to append to the analysis results field to enter an
appending file suffix. (Figure 3.15)
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Your currently displayed Result Name Files are appended (in real-time) as you type in
your suffix.
Figure 3.15 Adding a suffix
NOTE: If you are saving the same OSCHP file into the same output file folder that contains
your originally run OSCHP file with an identical suffix, a “2” is automatically added to the
filename to differentiate the two runs of identical CEL file names.
Exporting Batch Analysis Files (Optional)
You can export the information shown in the AT, GC, and Results File Names fields to
Microsoft Excel as a tab-delimited file for review and/or further batch editing.
NOTE: Once an analysis is submitted, a tab delimited file containing the cel file selections
is automatically saved in your designated output folder.
To export your batch analysis files:
1. Click Export Batch File.
A File window appears.
2. Navigate to the location where you want to save the file.
3. Make sure the Files of type is set to Tab Delimited File(s), then click Save.
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Log File Collection
Do the following if you experience any issues or failures with your analysis:
1. Click the Utilities button (top right of the OncoScan Console window)
2. Click to select Log Collection.
The following window appears. (Figure 3.16)
Figure 3.16 Log Collection File window
3. Use OncoScan Console’s default location of C:\ or navigate to a folder location of your
choice.
4. Click Create New Folder, then enter a folder name for your log.
5. Click OK.
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The following window appears confirming your log file has been saved as a zip file.
Figure 3.17 Example: Zip file contents of a Log Collection
6. Click OK to close the window.
NOTE: The auto-generated log collection zip file contains the full contents of the folder
and all QC History log files found in the configured QC History File path. By default, the
zip file resides here: C:\ProgramData\Affymetrix\Oncoscan\log
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Viewing the Log Collection File
To view the log collection file:
1. Use Windows Explorer to navigate to the location.
(Example: C:\ProgramData\Affymetrix\OncoScan\log)
2. Locate the zip folder you created earlier, then double-click on it.
The folder opens.
3. Extract the zipped folder’s contents, as you normally would. (Figure 3.18)
Figure 3.18 Example: Zip file contents of a Log Collection
Log Rollover
When the software determines that the log file for the Analysis Workflow
(C:\ProgramData\Affymetrix\OncoScan\log\AnalysisWorkflow.log) has reached
a defined size (approximately 4MB), the following steps will be completed:
A sub-folder will be created in C:\ProgramData\Affymetrix\OncoScan\log called
'Log*' (the '*' denotes the current date and time).
A zip file called RolledLogFile*.zip is created in that folder. The '*' is the same date and
time used for the folder name. The files in the
C:\ProgramData\Affymetrix\OncoScan\log folder and all files found in the currently
selected QC History Log folder will be included in this zip file.
The Analysis Workflow files that are associated with analysis workflows that are no longer
active on the Dashboard will be deleted from:
C:\ProgramData\Affymetrix\OncoScan\log
A new AnalysisWorkflow.log file will be created here:
C:\ProgramData\Affymetrix\OncoScan\log
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Analysis Submission and QC Results
Submitting your Analysis Setup Information
To submit your analysis:
1. After the information in the Analysis Setup window/tab is complete, click Submit.
The Workflow dashboard tab appears and processing begins.
Workflow Dashboard
The OncoScan Console Analysis Workflow Dashboard uses a progress bar to track the
software’s ongoing analysis tasks, then delivers the results of analyses. (Figure 4.1)
Figure 4.1 .CEL file analysis inside the Workflow dashboard
To pause and restart a Workflow analysis in progress:
1. Click Pause to stop the Workflow that is in progress.
2. Click Resume to restart the Workflow analysis.
To abort the Workflow in progress:
1. Click Pause to stop the Workflow that is in progress.
1. Click the X (upper right corner) of the Workflow pane.
A warning message appears.
2. Click OK to acknowledge the message.
After analysis is complete, a Workflow completed successfully message appears.
(Figure 4.2)
Figure 4.2 Workflow Dashboard example with multiple Single Samples loaded
3. To view the results, click
.
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The results appear in the QC Results tab. (Figure 4.3)
Figure 4.3 QC Results window tab
4. Click the checkbox (far left) of each sample you want to include in your report or click
Select All to auto-select all your samples.
QC Results
Interpreting and Using QC Results
The column header contains the metric name and its threshold (if a threshold has
been defined). See Customizing QC Metrics and Thresholds on page 30 to add a
threshold.

A CEL file value that does not pass its threshold test is indicated with an orange
background, as shown in Figure 4.3.

The Threshold Test column displays Outside Bounds and is highlighted in orange if
any metric in the row fails its threshold test, as shown in Figure 4.3.

A column labeled Within Bounds (as shown in Figure 4.3) indicates that all metrics on
the row passed the threshold test (or did not have a threshold applied to them).

Customizing QC Metrics and Thresholds
To add or remove QC metrics or the threshold associated with the metric:
1. Click Edit or Create QC Settings.
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The following window appears: (Figure 4.4 on page 31)
Figure 4.4 Edit or create QC Results window tab
The existing QC Metric OncoScan.Default contains the main metrics used in determining
whether the array passes or not.
OncoScan.All contains additional algorithm metrics that can be used for advanced
troubleshooting.
To view the Thresholds included with OncoScan Console, see Appendix , Appendix: QC
Metrics - Definitions, on page 68.
OncoScan™ Console 1.1 User Manual
To change a metric:
1. Click the Threshold’s drop-down to select a different Threshold. (Figure 4.5):
Figure 4.5 Threshold drop-down menu
2. Click the Threshold Option’s drop-down, then click to select a different symbol.
(Figure 4.6):
Figure 4.6 Threshold Options drop-down menu
If desired select a threshold value to help determine a prior Out of bounds result.
3. Click inside the Threshold Value’s field.
4. A cursor appears.
5. Use the backspace key, then enter a new value. (Figure 4.7):
Figure 4.7 Threshold Value field
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To add a new a QC Metric(s):
1. Click Add Threshold.
A new Threshold is added to the table.
2. Click the Threshold’s drop-down menu to select your new threshold.
A new Threshold Option is added to the new row.
3. Click the Threshold Option’s drop-down, then click to select a symbol.
A text box for Threshold Value is added to the column.
4. Click inside the Threshold Value’s field.
A cursor appears.
5. Enter a new value
6. You must enter a filename unless you are editing (and plan to overwrite) a previous QC
Metric filename
7. Click Save.
Exporting the QC Results Table
To export your QC Results table:
1. Click Export QC Table to export all the data shown in the table (no checking of the
checboxes is required).
A File window appears.
2. Navigate to the location you want.
3. Enter a File Name or use the default QCMetrixTable.txt.
Make sure the Files of type is set to Tab Delimited File(s).
4. Click Save.
The tab-delimited text version of the QC results table is now saved for your records.
(Figure 4.8).
Figure 4.8 Exported as a tab-delimited text file
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Generating and Exporting Reports
To Generate and Export your Results File table(s) as a tab-delimited text file:
1. Click the checkbox next to the Results File(s) you want to generate a report for, or click
Select All.
2. Click
to display the report menu options.
CelPairCheck Report on page 34

Gene Report on page 37

Probe Level Data Report on page 41

Segment Data Report on page 44

Somatic Mutation Data Report on page 47

Export All Data on page 51

CelPairCheck Report
This report is based off the signature SNPs and indicates whether the cel files selected as
the AT and GC files were likely from the same sample and assigned to the correct channel.
Do the following to export a CelPairCheck Report (aka SignatureSNP Report):
1. Click Export CelPairCheck Report. (Figure 4.9)
Figure 4.9 Generate Report dropdown menu
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Your previously assigned Output folder file window appears. (Figure 4.10)
Figure 4.10 OncoScan Output folder window
If you have not yet assigned an output folder, see Assigning an Output Results Path on
page 11.
2. The default root filename is Result. Click inside the File Name field to enter a different
root filename, then click Save.
A progress bar appears while your report generates, followed by Figure 4.11.
Figure 4.11 CelPairCheck Report finished
successfully.
3. Click Yes.
The OncoScan Output folder window appears.
4. Locate the SignatureSNP Report text file, then open it in Microsoft Excel.
The following window appears. (Figure 4.12)
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Figure 4.12 SignatureSNP report
SignatureSNP Report Column Definitions
Filename
Name of the OSCHP file containing the data
CEL Filename
Name of cel file.
Channel
The Channel file from which the signal is measured. "A" is the AT CEL, "C" is the GC
CEL.
CelPairCheckStatus
CelPairCheck is a test that inspects each pair of intensity (*.cel) files to determine
whether the files have been properly paired and assigned to the correct channel. In
addition to accidental mispairing of intensity files while setting up the analysis, a
tracking problem during the assay may result in a sample being assigned to the wrong
GeneChip array. As a result CelPairCheck ignores file names, and instead inspects the
genotypes in the two intensity files to detect file mispairings. To learn more about
CelPairCheck Status, see page 77.
CelPairCheckCallRate
CelPairCheckCallRate is the percentage of signature SNPs that make a genotype call for
a given CEL file.
CelPairCompareRate
This metric is the percentage of signature SNP control markers whose genotypes are
compared between the AT and GC channels.
CelPairConcordance
This metric is the concordance of a set of signature SNP genotypes compared between
AT and GC CEL files. If CelPairCheck Compare Rate is high but CelPairCheck
Concordance is low, then CelPairCheck Status will report "PossibleCELmispair".
SIG_001..00N
Genotype for signature snp 1..n
OncoScan™ Console 1.1 User Manual
Gene Report
This report summarizes the copy number segments that overlap user defined regions of
interest (e.g., Genes) as defined in the selected BED file.
Do the following to export a Gene Report:
1. Click Export Gene Report. (Figure 4.13)
Figure 4.13 Generate Report drop-down menu
The following window appears. (Figure 4.14)
Figure 4.14 Select the BED file for the Gene Report window
2. Click to select the appropriate BED file, then click Open.
NOTE: As shown in Figure 4.14, the default OncoScan-specific BED file for the Gene
report is OncoScanGeneBoundaries.r1.bed. However, any BED file can be used to
generate the Gene Report on any regions of interest contained within the BED file.
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Your previously assigned Output folder file window appears. (Figure 4.10)
Figure 4.15 OncoScan Output folder window
If you have not yet assigned an output folder, see Assigning an Output Results Path on
page 11.
3. The default root filename is Result. Click inside the File Name field to enter a different
root filename, then click Save.
A progress bar appears while your report generates, followed by Figure 4.11.
Figure 4.16 Gene Report finished successfully.
4. Click Yes.
The OncoScan Output folder window appears.
5. Locate the Gene Report text file, then open it in Microsoft Excel.
The following window appears. (Figure 4.17)
OncoScan™ Console 1.1 User Manual
Figure 4.17 Gene Report report
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Gene Report Column Definitions
Filename
Name of the OSCHP file containing the data
Chromosome
Chromosome on which the probeset is located.
Start Position
Start position of gene or region as defined in the bed file.
End Position
End position of gene or region as defined in the bed file.
Genes
This column is populated from the name column of the bed file. In most cases, it will
contain gene names.
Threshold Test
Displays Outside Bounds if any of the QC metrics fail to meet a threshold test. For more
information on thresholds, see Customizing QC Metrics and Thresholds on page 30.
% Aberr.Cells
If % AC = 100%, we return “homogeneous” because it could be 100% normal or 100%
tumor. If % AC =NA, the percent aberrant cells could not be determined and TuScan
returns non-integer CN calls. This metric is an algorithmically determined estimate of
the % of aberrant cells in the sample.
TuScan Ploidy
TuScan Ploidy is the most likely ploidy state of the tumor before additional aberrations
occurred. Algorithmically it is the CN state of the markers identified by the algorithm
as normal diploid before %AC and ploidy are determined. When a high ploidy is
determined the "normal diploid" is deemed to correspond to a higher CN and the log2
ratio gets adjusted appropriately. If ploidy cannot be determined NA (Not Available) is
reported.
Low Diploid Flag
An essential part of the algorithm is the identification of “normal diploid” markers in
the cancer samples. This is particularly important in highly aberrated samples. The
normal diploid markers are used to calibrate the signals so that “normal diploid
markers” result in a log2 ratio of 0 (e.g. copy number 2). The algorithm might later
determine that the "normal diploid" markers identified really correspond to (for
example) CN=4. In this case the log2 ratio gets readjusted and TuScan ploidy will report
4. Occasionally (in about 2% of samples) the algorithm cannot identify a sufficient
number of “normal diploid” markers and no “normal diploid calibration occurs. This
event triggers “low diploid flag” = YES. In this case the user needs to carefully examine
the log2 ratios and verify if re-centering is necessary.
Median Log2 Ratio
Log2 Ratio is the log2 ratio of the normalized intensity of the sample over the
normalized intensity of a reference with further correction for sample specific
variation. The Median Log2 Ratio is computed for each segment.
Median BAF
B-allele frequency (BAF) is (Signal (B)/{Signal(A) + Signal(B), where signal (A) is the
signal from the AT chip and signal (B) is the signal from the G/C chip. Median BAF is
reported for each segment and is the median BAF of the markers identified as
heterozygous, after mirroring any marker BAFs above 0.5 to the equivalent value
below 0.5. If the number of heterozygous markers in the segment is below 10 or the
percent of homozygous markers is above 85% no value is reported,
State
This is a comma separated list of the copy number state of the segments that overlap
the gene or region.
LOH
Flag to indicate whether the gene or region is in a Loss of Heterozygosity region (0=No,
1=Yes).
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Probe Level Data Report
This report contains base level data for each probeset including the log2ratio and BAF
values.
Do the following to export a Probe Level Data Report:
1. Click Export Probe Level Data. (Figure 4.18)
Figure 4.18 Generate Report
drop-down menu
Your previously assigned Output folder file window appears. (Figure 4.19)
Figure 4.19 OncoScan Output folder window
If you have not yet assigned an output folder, see Assigning an Output Results Path on
page 11.
2. The default root filename is Result. Click inside the File Name field to enter a different
root filename, then click Save.
OncoScan™ Console 1.1 User Manual
A progress bar appears while your report generates, followed by Figure 4.20.
Figure 4.20 Probe Level Data Report finished
successfully.
3. Click Yes.
The OncoScan Output folder window appears.
4. Locate the Probe Level Report text file, then open it in Microsoft Excel.
The following window appears. (Figure 4.21)
Figure 4.21 Probe Level report
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Probe Level Data Report Column Definitions
ProbeSet Name
Affymetrix identifier for the marker.
Chromosome
Chromosome on which the probeset is located.
Position
Chromosomal position of the probeset.
Log2 Ratio
Per marker Log2 Ratio of normalized intensity with respect to a reference, with further
correction for sample specific variation.
WeightedLog2Ratio
Contains the Log2 Ratios processed through a Bayes wavelet shrinkage estimator.
AllelicDifference
Allele difference is computed based on differencing A signal and B signal, then
standardizing based on reference file information.
NormalDiploid
Identifies the markers initially designated to be in a normal diploid region Used to
select the subset of data for generating the "sample sketch", which is used to quantile
normalize the raw intensities prior to further analysis. When the number of Normal
Diploid identified falls below a threshold, the "Low Diploid Flag" is set to "yes" and
the sample is normalized using all automsomal markers. As a result it is generally not
centered correctly, e.g. markers with log2 ratio of 0 may not correspond to CN=2.
BAF
BAF is (Signal (B)/{Signal(A) + Signal(B), where signal (A) is the signal from the AT chip
and signal (B) is the signal from the G/C chip.
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Segment Data Report
This report contains a list of all of the segments of normal and non-normal copy number
states and LOH found in the sample.
Do the following to export a Segment Data Report:
1. Click Export Segment Data. (Figure 4.22)
Figure 4.22 Generate Report
drop-down menu
Your previously assigned Output folder file window appears. (Figure 4.23)
Figure 4.23 OncoScan Output folder window
If you have not yet assigned an output folder, see Assigning an Output Results Path on
page 11.
2. The default root filename is Result. Click inside the File Name field to enter a different
root filename, then click Save.
OncoScan™ Console 1.1 User Manual
A progress bar appears while your report generates, followed by Figure 4.24.
Figure 4.24 Segment Data Report finished
successfully.
3. Click Yes.
The OncoScan Output folder window appears.
4. Locate the Segment Data Report text file, then open it in Microsoft Excel.
The following window appears. (Figure 4.25)
Figure 4.25 Segment Data report
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Segment Data Report Column Definitions
Segment ID
An Affymetrix identifier for the segment.
Chromosome
Chromosome on which the probeset is located.
Start Position
Start position of the segment.
End Position
End position of segment.
Marker Count
Number of markers in the segment.
Type
Indicates if the segment is a copy number segment or an LOH segment.
State
Indicates the copy number state of the segment for copy number segments or if the
segment contains LOH for LOH segments (0=No, 1 = Yes).
Median Log2 Ratio
Log2 Ratio is the log2 ratio of the normalized intensity of the sample over the
normalized intensity of a reference with further correction for sample specific
variation. The Median Log2 Ratio is computed for each segment.
Median BAF
B-allele frequency (BAF) is (Signal (B)/{Signal(A) + Signal(B), where signal (A) is the
signal from the AT chip and signal (B) is the signal from the G/C chip. Median BAF is
computed for each segment and is the median BAF of the markers identified as
heterozygous, after mirroring any marker BAFs above 0.5 to the equivalent value
below 0.5. If the number of heterozygous markers in the segment is below 10 or the
percent of homozygous markers is above 85% no value is reported.
% Aberr.Cells
If % AC = 100%, we return “homogeneous” because it could be 100% normal or 100%
tumor. If % AC =NA, the percent aberrant cells could not be determined and TuScan
returns non-integer CN calls. This metric is an algorithmically determined estimate of
the % of aberrant cells in the sample.
TuScan Ploidy
TuScan Ploidy is the most likely ploidy state of the tumor before additional aberrations
occurred. Algorithmically it is the CN state of the markers identified by the algorithm
as normal diploid before %AC and ploidy are determined. When a high ploidy is
determined the "normal diploid" is deemed to correspond to a higher CN and the log2
ratio gets adjusted appropriately. If ploidy cannot be determined NA (Not Available) is
reported.
Filename
Name of the OSCHP file containing the data
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Somatic Mutation Data Report
This report generates two (tab-delimited) text files; the somatic mutation file containing
the call for each somatic mutation in the sample, and the somatic mutation annotation file
containing annotation information for the somatic mutations assayed.
Do the following to export a Somatic Mutation Data Report:
1. Click Export Somatic Mutation Data. (Figure 4.26)
Figure 4.26 Generate Report drop-down
menu
Your previously assigned Output folder file window appears. (Figure 4.23)
Figure 4.27 OncoScan Output folder window
If you have not yet assigned an output folder, see Assigning an Output Results Path on
page 11.
2. The default root filename is Result. Click inside the File Name field to enter a different
root filename, then click Save.
NOTE: The Export Somatic Mutation Data report produces two separate (tab-delimited)
text files, as shown in the Files of type field. (Figure 4.27)
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A progress bar appears while your report generates, followed by Figure 4.24.
Figure 4.28 Somatic Mutation Data Report finished
successfully.
3. Click Yes.
The OncoScan Output folder window appears.
4. Locate the Somatic Mutation Data Report (*.somatic), then open it in Microsoft Excel.
The following window appears. (Figure 4.29)
Figure 4.29 Somatic (*.somatic) Data report
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Somatic (*.somatic) Data Report Column Definitions
Filename
Name of the OSCHP file containing the data
ProbeSetName
Name of the probeset.
MutCall
An indication of the whether the somatic mutation was decteded. A MutCall is
displayed as Undetected if the MutScore is below the Low Confidence threshold. A
MutCall is reported as HighConfidence if greater than or equal to the High Confidence
threshold. If the MutCall is equal to or greater than the Low Confidence threshold and
is less than the High Confidence threshold, the MutCall is reported as
LowerConfidence.
Note: MutCalls from"Outside Bounds" samples are not reliable.
MutScore
MutScore = (measured quantile normalized signal - median signal for this marker in the
reference model file) / (95th percentile signal for this marker in the reference model
file - median signal for this marker in the reference model file).
MutThreshHigh
High confidence MutScore threshold. Measurements equal to or greater than this
threshold are called "High confidence," describing the likelihood that the mutation is
present.
MutThreshLow
Lower confidence MutScore threshold. Measurements with a MutScore below this
value are called "Undetected". Measurements equal to or greater than this threshold
but less than the High Threshold are called "Lower confidence," describing the
likelihood that the mutation is present.
5. From your OncoScan Output folder, locate the second Somatic Mutation Data Report
(*.somaticannotation) text file, then open it in Microsoft Excel.
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The following window appears. (Figure 4.30)
Figure 4.30 Somatic (*.somaticannotation) report
Somatic (*.somaticmutation) Data Report Column Definitions
ProbeSetName
Name of the probeset.
chr_id
Chromosome on which the somatic mutation is found.
start
Start position of the somatic mutation.
stop
End position of the somatic mutation.
probeset_type
Indicates if the probeset is used for Somatic Mutation analysis (SOM).
tag_id
An Affymetrix identifier for the tag associated with the particular probeset.
common_id
Abbreviated description of the mutations to which this ProbeSet is known to respond.
The name has the form [Gene]:[amino acid change for mutation]:[cDNA change for
mutation]. In the event that the ProbeSet cannot differentiate among multiple
mutations to which it can respond, the slash (/) delimits the multiple known mutations.
cosmic_id
The identifier of the mutation as listed in the COSMIC database, which is a catalogue
of somatic mutations in cancer. More information on these mutations can be found at:
http://cancer.sanger.ac.uk
channel
The Channel file from which the signal is measured. "A" is the AT CEL, "C" is the GC
CEL.
OncoScan™ Console 1.1 User Manual
Export All Data
Use this option to generate all the reports (described in detail above) simultaneously.
1. Click Export All Data. (Figure 4.31)
Figure 4.31 Generate Report drop-down menu
The following window appears. (Figure 4.32)
Figure 4.32 Select the BED file for the Gene Report window
2. Click to select the appropriate BED file, then click Open.
NOTE: As shown in Figure 4.32, the default OncoScan-specific BED file for the Gene
report is OncoScanGeneBoundaries.r1.bed. However, any BED file can be used to
generate the Gene Report on any regions of interest contained within the BED file.
Your previously assigned Output folder file window appears. If you have not yet
assigned an output folder, see Assigning an Output Results Path on page 11.
NOTE: The default root filename is Result. Click inside the File Name field to enter a
different root filename.
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3. Enter a Root File Name for your text (tab-delimited) Export All Data file, then click
Save.
A progress bar appears while your report generates, followed by Figure 4.33.
Figure 4.33 Export All Data finished successfully.
4. Click Yes.
Your OncoScan Output folder window appears and shows all the reports generated
from the Export All option. (Figure 4.34)
Figure 4.34 Output folder window
5. Open the text file report you want to view using Microsoft Excel.
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Matched Normal Analysis Setup
To setup a Standard Analysis (See Standard Analysis Setup, on page 15)
Setting Up a Matched Normal Analysis
Figure 5.1 Matched Normal Analysis Setup window/tab - Overview
As long as your library file folder contains the necessary analysis files for the array, your
configuration paths are established and your Array Information fields auto-populate.
(Figure 5.2)
Figure 5.2 Matched Normal Analysis Configuration
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Confirming or Changing Array Information
1. From the Select array type drop-down list, select OncoScan.
NOTE: The Select array type drop-down list includes only the array types from the library
(analysis) files that have been downloaded from NetAffx or copied from the Library
package provided in the OncoScan installation package.
IMPORTANT: After adding new library files to the library file folder, always close and relaunch OncoScan Console to ensure the newly added files are recognized by the software.
2. From the Select analysis workflow drop-down list, click to select FFPE Analysis
including Matched Normal NA33.
Other available Analysis Workflow options are:
Ref103 Analysis NA33 - Use this workflow with cell line DNA.

FFPE Analysis NA33 - Use this workflow for a standard analysis. (See Setting

Up a Standard Analysis, on page 15)
3. Enter a Workflow name (optional). By default, the Set workflow name is Workflow.
Click
(upper right) to enter a different workflow name.
TIP: Customizing a Workflow name can be a useful tool in keeping track of analysis
workflows as all the related output files (outside of the OSCHP file) are pre-fixed with this
workflow name.
The Annotation file is automatically selected for you and is based on your selected
reference model file. (Example: OncoScan.na33.v1.annot.db))
NOTE: The Annotation to be used for analysis field is auto-populated based on your Ref
Model file selection. The analysis is not be permitted to run if the appropriate annotation
file is not available in your Library folder.
4. Select a Somatic mutation reference model file. By default, it is set to
OncoScan.na33.v1.SOM_REF_MODEL. If you created your own reference model file,
click the drop-down list to select your .SOM_REF_MODEL.
5. Confirm the displayed Somatic mutation threshold file to be used is correct. If you need
to change it, click the Browse button, navigate to the appropriate threshold .txt file,
then click OK.
IMPORTANT: If the Reference Model File and Somatic mutation Reference Model File
were created independently of each other, a warning message appears after you click
Submit (to start the Workflow Analysis process). Click OK to acknowledge the message.
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Adding CEL Files to Analyze
You can manually add CEL files or import them as a tab-delimited text file.
Manually Adding CEL Files to Analyze
To add batch-edited CEL files, see Importing CEL Files Using Batch Import, on page 58.
To manually add CEL files:
1. At the Select the intensity (CEL) file(s) to analyze pane, click the Add CEL files drop-
down.
2. Click Tumor AT Channel.
The CEL file window appears. (Figure 5.3)
NOTE: Immediately after a CEL file has been submitted for analysis (within the OncoScan
Console software), it goes into a locked state and cannot be used for other analyses until
the current analysis is complete.
Locked CEL file.
Figure 5.3 CEL file folder -EXAMPLE
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3. Click any header to sort your files or click the Files of type drop-down to filter your CEL
files by AT Channel, as shown in Figure 5.4.
Figure 5.4 Files of type drop-down list
4. Single click, Ctrl click, or Shift click (to select multiple Tumor AT Channel files).
IMPORTANT: Affymetrix recommends using an “A” or “C” as the last character to
designate the channel in the CEL file naming convention. Example: “_AS_05A.CEL” is an
AT Channel file, while “_AS_05C.CEL” is a GC Channel file. See Figure 5.3.
5. Click Open.
The Tumor AT Channel fields are now populated. (Figure 5.5)
Figure 5.5 Tumor AT Channel file list
6. Click the Add CEL files drop-down.
7. Click Tumor GC Channel. The CEL file window appears. (Figure 5.3 on page 55)
8. Single click, Ctrl click, or Shift click (to select multiple Tumor GC Channel files).
9. Click Open.
The Tumor GC Channel fields are now populated. (Figure 5.6)
Figure 5.6 Tumor GC Channel file list
10. Click the Add CEL files drop-down.
11. Click Normal AT Channel. The CEL file window appears. (Figure 5.3)
12. Single click, Ctrl click, or Shift click (to select multiple Normal AT Channel files).
13. Click Open.
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The Normal AT Channel fields are now populated. (Figure 5.7)
Figure 5.7 Normal AT Channel file list
14. Click the Add CEL files drop-down.
15. Click Normal GC Channel. The CEL file window appears. (Figure 5.3)
16. Single click, Ctrl click, or Shift click (to select multiple Normal GC Channel files).
17. Click Open.
The Normal GC Channel fields are now populated. (Figure 5.8)
Figure 5.8 Normal GC Channel file list
CEL File Displaying Options (Optional)
The File Name drop-down list (Figure 5.9) is dynamically populated and based on what
attributes are populated in the ARR file.
To use this display option, you must:
1. Provide the appropriate attributes at the time of sample registration in AGCC.
2. The ARR files must reside in the same folder as the CEL files.
Figure 5.9 EXAMPLE: File Name
Display Choices
To see “channel” (as an option in the drop down), you must use a template (or the
OncoScan template provided in the library files) that contains a “channel” attribute. The
resulting ARR file must also reside in the same folder as the CEL files you are analyzing.
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You can display one of the attributes from the ARR file in the table. For example,
“Channel” can be chosen (Figure 5.9) to confirm the assignment of a CEL file to its
appropriate channel.
To select a FIle Name display attribute:
1. Click the File Name drop-down button, then click to select the attribute you want
displayed along with your CEL file names.
The two examples (Figure 5.10 and Figure 5.11) show how the table appears with the
display set to Filename, then to Channel.
Figure 5.10 Table with Filename displayed
Figure 5.11 Table with Channel displayed
Importing CEL Files Using Batch Import
OncoScan Console allows import of CEL files using a batch file. The batch file must be saved
as a text (Tab-delimited) format and include the full directory path for your CEL files (as
shown in Figure 5.12).
TIP: The resulting OSCHP files are saved to your output path location, therefore it is not
necessary to include a path under RESULT. Simply enter the desired results filename in this
column.
The format for this tab-delimited file is 5 columns (A,B, C, D, and E) with the headers:
ATCHANNELCEL

GCCHANNELCEL

ATChannelMatchedNormalCel

GCChannelMatchedNormalCel

RESULT

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You must provide the full path to the CEL files for each Channel column.
(Example: C:\Desktop\OncoScan\Data\Sample1.cel)
Figure 5.12 List from Windows Excel
1. Click Import Batch File
A File window appears.
2. Navigate to your text (tab delimited) file location, then click on the file you want to
import.
IMPORTANT: The Microsoft Excel application must be closed before you import (click
Open).
3. Click Open.
The Tumor AT, Tumor GC, Normal AT, Normal GC and Result File Name fields are now
populated. (Figure 5.13)
Figure 5.13 Tab-delimited text file imported into OncoScan Console
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Generating Result File Names
Results File Names can either be entered in manually or OncoScan Console can generate
them automatically.
NOTE: If you use the suffix option (Selecting a Suffix to Append to the Analysis Results,
on page 62) and enter your Result File Names manually, your assigned suffix appears in
the Results File Name column.
If you auto-generate your Results File Names, your assigned suffix appears in the Results
File Name column, but it does get added to your final OSCHP file name(s).
To manually enter a Results File Name:
1. Single-click inside the appropriate Results Name File field to produce a cursor, then type
in the file name you want.
To auto-generate a suggested Result File Name:
NOTE: During the Result File Name auto-generation process, the file names are compared
to identify their common root name for use as a results file name. Generally, the last 5
characters of each CEL file name are ignored, then the remaining root names of the AT
and GC file names are compared. If the root names of the AT and GC channel match, then
the root name is used in the Results File Name field. The one exception is if your array
name “_(OncoScan)” is appended to the file name during registration in Affymetrix
GeneChip Command Console (AGCC). In this case, the “_(OncoScan)” is ignored during the
comparison, but then added back in the Results File Name field.
1. After the 4 Channel lists are populated, click the Result File Names drop-down, then
select Auto Generate Output Name.
2. The Result File Name column is now populated with suggested filenames for each
pairing. (Figure 5.14)
Figure 5.14 Matched Normal Results File Name list
Common root names should be consistent all the way up to the last character of the CEL
file name prior to the .cel extension. If there is a paired file mis-match, the Results File
Name appears as Output1. (Figure 5.15)
Figure 5.15 Result File Name “Output”
If Output1 or subsequent Outputs (Output 2, Output 3...) appear, investigate the validity
of your original pairing. See Correcting Mismatched CEL File Pairings, on page 61.
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To edit an auto-generated Result File Name:
1. Click on the Result File name you want to edit.
2. After the cursor appears, edit the filename as you normally would.
3. Click outside the row to save your edit.
To clear the entire Result File Name column:
1. Click the Result File Names drop-down button, then select Clear Column.
The column is now cleared and ready for new Result File Name entries.
Correcting Mismatched CEL File Pairings
If there is a paired file mismatch, the Results File Name appears as Output1, Output2,
Output3, etc.
A paired file mismatch is most likely caused by an incorrect CEL filename pairing and not a
mismatch of your native CEL files.
A simple way to correct mismatches is to sort the AT and GC columns so that files with the
same root names are next to each other.
TIP: Common root names should be consistent all the way up to the last character of the
cel file name prior to the .cel extension. Affymetrix recommends using an “A” or “C” as
the last character to designate the channel in the CEL file naming convention. Example:
“_AS_05A.CEL” is an AT Channel file, while “_AS_05C.CEL” is a GC Channel file.
Using the Sorting Features
To sort an individual column:
1. Click on a Channel header.
The column is now sorted in an ascending order.
2. Click on the Channel header again to reverse the sorting order.
To sort all the columns simultaneously:
1. Click Sort All.
The file contents of the 4 columns are now sorted together in an ascending order.
2. Click Sort All again.
The file contents of the 4 columns are now sorted together in a descending order.
To swap CEL files between columns:
1. Click and drag a column CEL entry onto another column CEL entry, then release the
mouse button.
The CEL entries have now swapped column positions.
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To reorder the CEL files in a column:
1. Click and drag a CEL file to another position within the column, then release the mouse
button.
The CEL file is now at its new position.
To add a cell to a column:
1. Click and drag a column cell to the top or bottom border line of a neighboring cell, then
release the mouse button.
Generating a Result File Name after Sorting
1. After all your columns are properly sorted, click the Result File Names drop-down,
then select Auto Generate Output Names.
The Result File Name column is now populated with suggested filenames for each
pairing.
NOTE: An OSCHP file is created for each pair. The tumor will have “_T” and the normal
will have “_N” appended to its root name.
If OncoScan Console detects an inconsistency between the AT and GC file names to be
paired, a Result File Name labeled, “Output n” reappears.
IMPORTANT: Confirm that both columns are sorted in the same direction. If they are,
examine the files and confirm they are paired correctly. The file names (excluding the last
character before the .CEL) MUST match exactly.
Repeat the sorting steps above, then try to Auto Generate Output Names again until a
successful Result File Name(s) appears.
Setting your Output Information Location (Optional)
The Output result information path (lower left) is retained from your initial setup.
To select a different folder to store your results:
1. Click the browse button, then navigate to the folder you want. If you want to change
the default folder, see Assigning an Output Results Path, on page 11.
Selecting a Suffix to Append to the Analysis Results
You can append a suffix at the end of all your Results File Names. This is useful when
tracking versions of the analysis files used to generate the resulting OSCHP files.
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To use an appending suffix:
1. Click inside the Select a suffix to append to the analysis results field to enter an
appending file suffix. (Figure 5.16)
Your currently displayed Result Name Files are appended (in real-time) as you type in
your suffix.
Figure 5.16 Adding a suffix
NOTE: If you are saving the same OSCHP file into the same output file folder that contains
your originally run OSCHP file with an identical suffix, a “2” is automatically added to the
filename to differentiate the 2 runs of identical CEL file names.
Exporting Batch Analysis Files (Optional)
You can export the information shown in the Tumor and Normal AT and GC Channels and
Results File names fields to Microsoft Excel as a tab-delimited file for review and/or further
batch editing.
NOTE: Once an analysis is submitted, a tab delimited file containing the cel file selections
is automatically saved in your designated output folder.
To export your batch analysis files:
1. Click Export Batch File.
A File window appears.
2. Navigate to the location where you want to save the file.
3. Make sure the Files of type is set to Tab Delimited File(s), then click Save.
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Log File Collection
Do the following if you experience any issues or failures with your analysis:
1. Click the Utilities button (top right of the OncoScan Console window)
2. Click to select Log Collection.
The following window appears. (Figure 5.17)
Figure 5.17 Log Collection File window
3. Use OncoScan Console’s default location of C:\ or navigate to a folder location of your
choice.
4. Click Create New Folder, then enter a folder name for your log.
5. Click OK.
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The following window appears confirming your log file has been saved as a zip file.
Figure 5.18 Example: Zip file contents of a Log Collection
6. Click OK to close the window.
NOTE: The auto-generated log collection zip file contains the full contents of the folder
and all QC History log files found in the configured QC History File path. By default, the
zip file resides here: C:\ProgramData\Affymetrix\Oncoscan\log
Viewing the Log Collection File
To view the log collection file:
1. Use Windows Explorer to navigate to the location.
(Example: C:\ProgramData\Affymetrix\OncoScan\log)
2. Locate the zip folder you created earlier, then double-click on it.
The folder opens.
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3. Extract the zipped folder’s contents, as you normally would. (Figure 5.19)
Figure 5.19 Example: Zip file contents of a Log Collection
Log Rollover
When the software determines that the log file for the Analysis Workflow
(C:\ProgramData\Affymetrix\OncoScan\log\AnalysisWorkflow.log) has reached
a defined size (approximately 4MB), the following steps will be completed:
A sub-folder will be created in C:\ProgramData\Affymetrix\OncoScan\log called
'Log*' (the '*' denotes the current date and time).
A zip file called RolledLogFile*.zip is created in that folder. The '*' is the same date and
time used for the folder name. The files in the
C:\ProgramData\Affymetrix\OncoScan\log folder and all files found in the currently
selected QC History Log folder will be included in this zip file.
The Analysis Workflow files that are associated with analysis workflows that are no longer
active on the Dashboard will be deleted from:
C:\ProgramData\Affymetrix\OncoScan\log
A new AnalysisWorkflow.log file will be created here:
C:\ProgramData\Affymetrix\OncoScan\log
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Appendix: Custom Reference Files
Creating your own Reference File
To create a reference file:
IMPORTANT: When creating an OncoScan FFPE Assay reference file, you must use a
minimum of 6 CEL files (including 1 male and 1 female). It is recommended that you select
at least 44 normal samples of good quality. Ideally, your male and female samples should
be of equal numbers (22 and 22).
1. From the Analysis Setup tab Select array type drop-down list, click to select an array type
(Example: OncoScan) (Figure A.1)
Figure A.1 Reference File creation example
NOTE: The Select array type drop-down list includes only the array types for which library
(analysis) files have been downloaded from NetAffx or copied from the Library package
provided with the installation.
2. From the Select analysis workflow drop-down list, click to select Reference
Generation NA33.
3. By default, the Set workflow name is Workflow. Click
(upper right) to enter
a different workflow name.
4. Select the annotation File for this analysis to be used for analysis. (Example:
OncoScan.na33.v1.annot.db)
After you create your reference file, navigate to your library file folder, then open the
Signature SNPs Report text file. Confirm that the report shows that your sample pairings
and channel IDs are correct.
If they are not, you must adjust the pairings and channel data in OncoScan Console’s
Select the intensity (CEL) file(s) to analyze table (see Correcting Mismatched CEL File
Pairings on page 18), then re-run the reference file.
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Appendix: QC Metrics - Definitions
Array Data QC Metrics (Overview)
This section provides a high level overview of the key QC metrics used with OncoScan
Console.
MAPD (Median of the Absolute Values of all Pairwise Differences)
MAPD is a global measure of the variation of all microarray probes across the genome. It
represents the median of the distribution of changes in log2 ratio between adjacent
probes. Since it measures differences between adjacent probes, it is a measure of shortrange noise in the microarray data. Lower MAPD values are better. For more information,
see Array Data QC Metrics (Detailed Descriptions) on page 73.
ndSNPQC (SNP Quality Control of Normal Diploid Markers)
The metric, SNPQC is a measure of how well genotype alleles are resolved in the microarray
data. ndSNPQC is the same metric but only applied to normal diploid markers (that is those
that have been determined to have Copy Number =2 in the sample). Larger ndSNPQC
values are better. For more information, see ndSNPQC on page 75.
SNP QC Type (SNP Quality Control Type)
If SNP QC Type is ND, metrics like ndSNPQC, ndWavinessSD, ndSNR_AT, ndSNR_GC,
ndRawSNPQC are all based on the performance of the normal diploid marker subset
counted by ndCount. If SNP QC Type is non ND, then ndSNPQC and ndRawSNPQC are
computed based on a preselected set of 10,000 autosomal markers, while ndWavinessSD
are computed on all autosomal markers.
CelPairCheck Status
CelPairCheck is a test that inspects each pair of intensity (*.cel) files to determine whether
the files have been properly paired and assigned to the correct channel. In addition to
accidental mispairing of intensity files while setting up the analysis, a tracking problem
during the assay may result in a sample being assigned to the wrong GeneChip array. As a
result CelPairCheck ignores file names, and instead inspects the genotypes in the two
intensity files to detect file mispairings. If the CelPairCheck Status is not Pass, see
CelPairCheckStatus on page 77.
CelPairCheck Compare Rate
This metric is the percentage of signature SNP control markers whose genotypes are
compared between the AT and GC channels. The Compare Rate needs to be above a
minimum in order for CelPairCheck to determine whether the AT and GC CEL files belong
to the same individual.
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CelPairCheck Concordance
This metric is the concordance of a set of signature SNP genotypes compared between AT
and GC CEL files. If CelPairCheck Compare Rate is high but CelPairCheck Concordance is
low, then CelPairCheck Status will report "PossibleCELmispair".
ndWavinessSD (Normal Diploid Waviness Standard Deviation)
ndWavinessSD is a global measure of variation of microarray probes that is insensitive to
short-range variation and focuses on long-range variation. ndWavinessSD is computed on
normal diploid markers.
ndWavinessSD should be used along with Low Diploid Flag, ndCount (the actual number
of diploid markers identified) BAFs, and log2 ratio to assess if the log2 ratio is centered
correctly. ndWavinessSD can thus help assess if log2 ratios need to be re-centered.
In addition when ndWavinessSD is high, the log2 ratios should be examined for clear
breakpoints as opposed to a gradual drift of the log2 ratio. When the latter is observed
small aberrations should be examined carefully. When breakpoints are sharp and the
ndCount is large a high ndWavinessSD can be ignored.
Y Gender Call
Gender call determined by examining signal on the Y Chromosome.
ndCount
ndCount reports how many normal diploid markers were identified. When ndCount falls
below a minimum, (typically 2000), the Low Diploid Flag is triggered. ndCount > 10,000 is
desired for maximum confidence in ndSNPQC and maximum confidence in the centering of
the log2 ratios.
Low Diploid Flag
An essential part of the algorithm is the identification of “normal diploid” markers in the
cancer samples. This is particularly important in highly aberrated samples. The normal
diploid markers are used to calibrate the signals so that “normal diploid markers” result in
a log2 ratio of 0 (e.g. copy number 2). The algorithm might later determine that the
"normal diploid" markers identified really correspond to (for example) CN=4. In this case
the log2 ratio gets readjusted and TuScan ploidy will report 4. Occasionally (in about 2%
of samples) the algorithm cannot identify a sufficient number of “normal diploid” markers
and no “normal diploid calibration occurs. This event triggers “low diploid flag” = YES. In
this case the user needs to carefully examine the log2 ratios and verify if re-centering is
necessary.
ACDC (Aberrant Cell-Derived Copy Number)
Indicates whether the algorithm was able to compute the % Aberr. Cells and measure the
Copy Number in the tumor cells only. “ACDC= No” means Copy Number was calculated as
an average CN across all cells, “ACDC=Yes” means that Copy Number was calculated only
for the tumor cells.
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%Aberr. Cells
Algorithmic estimation of the percent of aberrant cells (%AC) in a sample.
When %AC is “NA” it means that the % aberrant cells could not be estimated because the
percent is either too low or the sample is heterogeneous and hence is composed of several
types of aberrated cells, or the percent varies from one genomic location to another.
When %AC is not NA, (example 60%), it means that across all aberrations, ~60% of the cells
were aberrated and contributed to the elevated (or reduced) Copy number.
TuScan Ploidy
TuScan Ploidy is the most likely ploidy state of the tumor before additional aberrations
occurred. Algorithmically it is the CN state of the markers identified by the algorithm as
normal diploid before %AC and ploidy are determined. When a high ploidy is determined
the "normal diploid" is deemed to correspond to a higher CN and the log2 ratio gets
adjusted appropriately. If ploidy cannot be determined NA (Not Available) is reported.
Offset Flag
If the Offset Flag is Yes, the TuScan results required an adjustment to the log2 ratios, as
explained under TuScan Ploidy. As a result, allelic peaks, and smooth signal values get
rescaled accordingly.
L2R Offset
L2R Offset is the constant added to the log2 ratios, when the Offset Flag = Yes.
Low % Aberrant Cell nGoF
Goodness of fit (GoF) is evaluated when TuScan tries to explain the CN changes by assuming
a two state mixture model of normal and aberrant cells. A large nGoF value indicates an
estimation of the % aberrant cells cannot be calculated, due to the aberrant cell fraction
being below TuScan detection limits.
Hyb Control Intensity_AT
Geometric mean of three array hybridization controls in the AT channel. An unusually low
value indicates a problem at the array hybridization and/or washing step.
Hyb Control Intensity_GC
Geometric mean of three array hybridization controls in the GC channel. An unusually low
value indicates a problem at the array hybridization and/or washing step.
Q3 Raw Intensity_AT
The 75th percentile of the raw intensity of the AT channel.
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Q3 Raw Intensity_GC
The 75th percentile of the raw intensity of the GC channel.
AGR_AT
Antigenomic ratio in the AT channel. Measures non-specific binding to array features in
that channel. Smaller values are better.
AGR_GC
Antigenomic ratio in the GC channel. Measures non-specific binding to array features in
that channel. Smaller values are better.
ndSNR_AT
This metric is the Signal/Noise Ratio of normal diploid markers in the AT channel. The Signal
is the 65th percentile of the measurements used for copy number analysis, minus the
median of the antigenomic features. The Noise is the standard deviation of the weakest
15% of measurements used for copy number analysis. A low value indicates poor data
quality from the AT CEL file.
ndSNR_GC
This metric is the Signal/Noise Ratio of normal diploid markers in the GC channel. The
Signal is the 65th percentile of the measurements used for copy number analysis, minus the
median of the antigenomic features. The Noise is the standard deviation of the weakest
15% of measurements used for copy number analysis. A low value indicates poor data
quality from the GC CEL file.
ndRawSNPQC
Like ndSNPQC, but calculated on raw intensities.
Call Rate
The percentage of snps for which a genotype could be determined - divided by the total
number of snps. The higher the Call Rate, the better. Call rate is based on genotype calls
using an algorithm which assumes all autosomal markers are normal diploid. Therefore,
this metric is more relevant for normal samples than for aberrated tumor samples.
Matched Normal Compare Rate
If doing FFPE analysis including matched normal (where you supply four CEL files per row
during analysis setup), this metric reports the percentage of markers identified as normal
diploid in both the resulting Tumor OSCHP file, and its matched Normal OSCHP file. If this
metric is unusually low, then the results of the companion metric Matched Normal
Concordance may be unreliable.
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Matched Normal Concordance
If doing FFPE analysis including matched normal (where you supply four CEL files per row
during analysis setup), this metric reports the genotype concordance of the normal diploid
markers in common between the Tumor OSCHP file and its matched Normal OSCHP file. If
the Matched Normal Compare Rate is reasonably high but Matched Normal Concordance
is low, then the normal CEL files you paired with the tumor CEL files may not belong to the
same individual. This metric is only reported for the Tumor OSCHP file.
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Array Data QC Metrics (Detailed Descriptions)
MAPD
For quality assessment purposes, we define metrics that assess whether the microarray data
is of sufficient quality to accurately assess copy number (CN) analysis. One of these metrics
is Median of the Absolute values of all Pairwise Differences (MAPD).
MAPD is defined as the Median of the Absolute values of all Pairwise Differences between
log2 ratios for a given chip. Each pair is defined as adjacent in terms of genomic distance,
with SNP markers and CN markers being treated equally. Hence, any two markers that are
adjacent on the genome are a pair. Except at the beginning and the end of a chromosome,
every marker belongs to two pairs, as shown in Figure 1.
Figure 1 Example: Two pairs to a marker
Formally, if xi: is the log2 ratio for marker i:
MAPD = median(|xi-1 – xi|, with i ordered by genomic position)
MAPD is a per-microarray estimate of variability, like standard deviation (SD) or
interquartile range (IQR). If the log2 ratios are distributed normally with a constant SD,
then MAPD/0.96 is equal to SD and MAPD*1.41 is equal to IQR. However, unlike SD or IQR,
using MAPD is robust against high biological variability in log2 ratios induced by conditions
such as cancer.
Variability in log2 ratios in a microarray arises from two distinct sources:
Intrinsic variability in the starting material, hybridization cocktail preparation,
microarray or scanner

Apparent variability induced by the fact that the reference may have systematic
differences from this microarray

Regardless of the source of the variability, increased variability decreases the quality of CN
calls.
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Effect of MAPD on Functional Performance
As a measure of performance, we measured copy number gains and loss using over 1000
cancer samples. Arrays with MAPD > 0.3 are out of bounds.
ndWaviness-SD
Waviness refers to an effect seen in all genomic microarrays (see Maroni et al. (2007)
Genome Biology 8:R228) where long-range variation is observed, often associated with
regional genomic differences like local GC-content changes.
This metric is only computed on markers that have been identified as Normal diploid by the
algorithm. When Low Diploid Flag = Yes, ndWaviness-SD is computed for all autosomal
markers. For most samples, a ndWaviness-SD value below 0.12 for OncoScan arrays
indicates that the long-range variation is within levels that can be accommodated by the
algorithms.
ndWavinessSD should be used along with LowDiploidFlag, ndCount (the actual number of
diploid markers identified) BAFs and log2 ratio to assess if the log2 ratio is centered
correctly. ndWavinessSD can thus help assess if log2 ratios need to be re-centered.
In addition when ndWavinessSD is high, the log2 ratios should be examined for clear
breakpoints as opposed to a gradual drift of the log2 ratio. When the latter is observed
small aberrations should be examined carefully. When breakpoints are sharp and the
ndCount is large a high ndWavinessSD can be ignored.
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ndSNPQC
The ndSNPQC metric estimates the distributions of homozygous AA, heterozygous AB and
homozygous BB alleles and calculates the distance between them. The better the
separation of these distributions, the better the ability to identify a genotype based on its
cluster position, as shown in Figure 2.
Figure 2 Example: Genotype based on its cluster position
SNPQC correlates well with genotype performance, as measured by Call Rate and
Concordance to published HapMap genotypes.
For OncoScan, we use ndSNPQC. This is computed only on the normal diploid markers to
assess the quality of the genotypes and the resulting quality of BAFs. When Low Diploid
Flag = Yes, ndSNPQC is computed for a preselected set of 10,000 autosomal markers. As a
result, values may fall out of bounds although the data quality is good. We therefore
recommend always examining the BAF visually when Low Diploid Flag = Yes and ndSNPQC
is out of bounds.
Effect of ndSNPQC on Functional Performance
ndSNPQC provides insight into the overall level of data quality from a SNP perspective. The
key consideration when evaluating the ndSNPQC value is to ensure the threshold is
exceeded. The quality of the SNP allele data is compromised, and is noisier and more
difficult to interpret when the ndSNPQC values are below the recommended acceptance
threshold as illustrated in the figure below.
When the ndSNPQC value is below 26, the noise within the array is higher than normal
which compromises the overall clarity of results. ndSNPQC values values above 26 have
good data quality and can be relied upon with regards to performance as shown in Figure
3. Sometimes ndSNPQC values can be as high as 35 (or higher) and provide even better
separation of BAFs.
OncoScan™ Console 1.1 User Manual
Figure 3 Examples: (TOP) Sample with Noisier BAFs. (BOTTOM) Sample with clearly delineated BAFs.
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CelPairCheckStatus
CelPairCheckStatus
message
Description
Pass
No intensity file pairing problem is detected.
PossibleCELmispair
Low genotype concordance exists between data in AT channel and
data in GC channel. This is consistent with the data in the 2 intensity
files originating from different individuals.
This message may also appear with poor data. Please review the channel-specific sample QC metrics to see if one or both CEL files has a
problem.
PossibleGCinATchannel
Data from the GC reaction appears to be assigned to the AT analysis
channel.
PossibleATinGCchannel
Data from the AT reaction appears to be assigned to the GC analysis
channel.
Warn
Cannot determine whether the two intensity files belong to the same
individual.
This message may appear with poor data, or with good data where
too many of the signature SNPs used for CelPairCheck are in chromosome regions with copy number aberrations.
Most of the probes in the OncoScan assay are designed so that each OncoScan array detects
only one of the two alleles for a specific SNP, and so both arrays are needed to measure a
full genotype call for that SNP. However a set of control probes are included so that both
alleles for a set of “Signature SNPs” can be measured on each array independently.
The pattern of genotype calls for these Signature SNPs is compared between the two
intensity files. As the signature SNPs are a set of high minor allele frequency SNPs, under
normal assay conditions it is very unlikely that two unrelated tested samples will have the
same pattern of Signature SNP genotype calls. Therefore, if the concordance of genotype
calls is unusually low between a given pair of intensity files, the CelPairCheck Status
message will be "PossibleCELmispair." The concordance check is only done if there are
enough Signature SNPs reporting a call in both files. If this is not the case, CelPairCheck
Status reports “Warn.”
Signature SNP probes are designed to be channel-specific. When an intensity file is
assigned to the AT channel, only the Signature SNP probes designed to respond in the
assay's AT reaction well are expected to report genotypes. In the case of GC data being
assigned to the AT channel for analysis, a low call rate will result in the CelPairCheck Status
message “PossibleGCinATchannel.” If AT data were assigned to the GC channel, the
message would be "PossibleATinGCchannel."
If you need additional help fixing the problems detected by CelPairCheck Status, you can
review that Signature SNP metrics and genotypes for each intensity file in the
CelPairCheckReport file. This report is created along with result files (*.oschp) for each
analysis, or go to OncoScan Console’s QC results tab, click on the Generate Report
button, then select Export [CelPairCheck Report]. See Generating and Exporting Reports
on page 34 for more details.
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ndWavinessSD
For OncoScan we compute Waviness-SD on markers identified by the algorithm as “normal
diploid” markers. It is a QC metric that focuses on measuring long-range effects. As
described separately, MAPD is a metric that measures short-range variation, the variation
of adjacent probes. The long-range variation measurement is accomplished by calculating
the variation in log2 ratios across the whole genome and subtracting out the short-range
variation, specifically, for autosomal probes:
Define:
Xi as the log2 ratios of autosomal probes
And Zi as the variance between adjacent probes:
Zi = X2i+1 - X2i
Waviness-SD is the total variance (Xi) minus the local variance (Zi):
Waviness-SD = sqrt(Var(Xi)-Var(Zi)/2)
% Aberrant Cells
If % AC = 100%, we return “homogeneous” because it could be 100% normal or 100%
tumor. If % AC =NA, the percent aberrant cells could not be determined and TuScan
returns non-integer CN calls.
The cause could be:
1. Low % aberrant cells
2. Low number of diploid regions
3. Data Quality, in particular for all samples for which MAPD is above 0.4 or ndSNPQC falls
below 19.
4. Occasionally High/Low Ploidy
5. No Ploidy solution has a good fit to the data.
Low Diploid Flag
An essential part of the algorithm is the identification of “normal diploid” markers in the
cancer samples. This is particularly important in highly aberrated samples. The normal
diploid markers are used to calibrate the signals so that “normal diploid markers” result in
a log2 ratio of 0 (e.g. copy number 2). The algorithm might later determine that the
"normal diploid" markers identified really correspond to (for example) CN=4. In this case,
the log2 ratio gets readjusted and TuScan ploidy will report 4. Occasionally (in about 2%
of samples) the algorithm cannot identify a sufficient number of “normal diploid” markers
and no “normal diploid calibration occurs. This event triggers “low diploid flag” = YES. In
this case the user needs to carefully examine the log2 ratios and verify if re-centering is
necessary.
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Appendix: Algorithms
B-allele Frequencies
B-allele frequencies (BAFs) are a graphical way to show allelic imbalances. BAFs are also
used by the algorithm to derive the CN state. Conceptually for each marker we compute
(Signal (B)/{Signal(A) + Signal(B), where signal (A) is the signal from the AT chip and signal
(B) is the signal from the G/C chip. A homozygous BB SNP will therefore have a value of 1,
a homozygous AA SNP a value of zero and a AB SNP a value of 0,5. For SNPS with high
Minor allele frequency the BAFs present as three equally thick bands around these values
(0, 0.5, 1).
In regions of Loss of heterozygosity the only possible alleles are BB and AA and the middle
band (corresponding to AB) is missing.
In regions of Copy Gain the allelic balance is disrupted. With 3 copies it is not possible to
have an equal amount of A and B alleles, and the BAF bands are at 0, 1/3, 2/3 and 1.
In cancer samples the additional complication of normal contamination of the tumor cells
affects the allelic imbalance further.
When 60% of the cells have CN=3 and 40% of the cells have CN=2 the location of the BAF
bands is at (0.6*2+0.4)/(3*0.6+2*0.4) =0.615, while it is at 0.66 (or 2/3) when 100% of the
cells have CN=3. In general when the percent aberrated cells is p and the CN for these
aberrated cells is 3 then, the middle BAF band shifts up to (2p+(1-p))/(3p+2(1-p)) an down
to 1- (2p+(1-p))/(3p+2(1-p)).
LOH Algorithm
The LOH algorithm uses B-allele frequencies (BAFs) and log2 ratios to find long stretches
of homozygosity.
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TuScan Algorithm
The TuScan algorithm uses B-allele frequencies (BAFs) and log2 ratios to estimate the
ploidy and percentage of aberrant cells in the sample (%AC) which in turn are used to
calculate copy number calls (CN). The BAFs and log2 ratios contribute equally to CN
determination. TuScan first uses the BAFs and log2 ratio data to identify segments of equal
CN. Next TuScan uses the BAFs, log2ratios and segment data to find the combination of
%AC and ploidy that best fits the data. When TuScan can successfully determine %AC, the
algorithm assigns each aberrant segment an integer copy number representing the copy
number in the tumor portion of the sample. This is possible because CN is well
approximated by an integer when the tumor is nearly homogeneous. If the tumor is highly
heterogeneous (i.e., lacks a dominant clone), or contains a large amount of “normal” cells
%AC cannot be determined. In other words, if the percentage of aberrant cells
contributing to the various aberrations in the sample varies across all aberrations, %AC and
ploidy cannot be determined. When %AC cannot be determined, the segmentation
algorithm will still identify segments of equal CN, but the CN in just the aberrant cells
cannot be determined. In this case, TuScan bins the copy numbers and returns fractional
CN values in 1/3 increments (e.g., 2, 2.33, 2.66, 3 etc.). This fractional copy number is derived
from the normal contamination as well as the heterogeneous population of tumor cells;
therefore, the fractional CN calls represent the average CN observed for that segment.
Users should look at the value of %AC to determine whether the CN value represents the
CN in the tumor (%AC= number) or the average CN in the sample (%AC=NA). Tumor
heterogeneity also affects the interpretation of the CN number calls when %AC cannot be
determined. For example, a TuScan call of 2.33 can result from 40% of the aberrant cells
having 3 copies, 10% of aberrant cells having 5 copies, or a more complex heterogeneous
mixture of copy numbers. Since nearly every tumor sample will have some amount of
normal contamination combined with tumor heterogeneity it is not possible to predict how
often TuScan will be able to determine the %AC, it will vary depending on the sample.
OncoScan™ Console 1.1 User Manual
Figure 1 Example: TuScan results as displayed in OncoScan Nexus Express
81
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