How to use the BeadArray Controls Reporter (BACR) to quickly analyze Infinium MethylationEPIC Array

The BeadArray Controls Reporter (BACR) software can be used to quickly analyze the built-in controls on Infinium Methylation Arrays, including Infinium MethylationEPIC v2.

Why use BeadArray Controls Reporter?

• Computationally faster than making a GenomeStudio 2011.1 project to analyze data,

• Helpful for large datasets that may be analyzed with third-party software.

• Automatically compares expected positive signal with expected negative signal to remove background and flags controls that deviate from the expected relative intensity patterns with yellow highlighting.

• Populates a spreadsheet with results, which is convenient for keeping records/documenting run performance data.

How to run an analysis of Methylation Controls with BACR

1. Ensure that data output files from scanner are labeled with 12-digit barcode of beadchip, and that these folders contain .idat files (Be sure not to use the DMAP folders that are also named by beadchip barcode; the contents of the folder should contain .idat files, not .gz DMAP files).

2. Make a folder on your computer that contains the following:

• all IDAT folders to be included in analysis

• .bpm manifest file

• .csv Sample sheet. See Sample Sheet Creation section from BACR Software Guide below for instructions on how to prepare sample sheet

Example folder structure for folder called 'Demo Data EPIC'

1. Open Bead Array Controls Reporter (link to download software is below) and click the 'Browse' button to assign the 'IDAT File Location'. The IDAT File Location is the parent folder containing all the required files listed above-- in this case 'Demo Data EPIC'

4. Select 'Options' > 'OutputDirectory' to configure where you would like to save the output file.

1. Click 'Start Analysis' to run the application.

2. Open the output file using Microsoft Excel or a similar spreadsheet program. The output file will be saved to the location you selected and will have the name of the parent folder with the word 'controls' appended to the end of the file name.

3. Interpret the results

• The products of the calculations performed by BACR will be flagged with yellow highlighting in the spreadsheet if the thresholds set in the BACR application are exceeded. It is recommended to conduct a more through evaluation of the controls using GenomeStudio 2011.1 Controls Dashboard visualization tool if values are flagged as yellow.

• In some cases, the thresholds (including the Background Correction Offset) may need to be adjusted slightly depending on the scanner that was used.

BACR Options - Customizing Threshold Settings

To adjust the thresholds used by BACR to flag calculation products as yellow, from the Options menu, select 'Set Thresholds'

** If the FFPE restore kit was used, set the Restoration Threshold to 1. Otherwise, leave at default setting (0). ** Only adjust Thresholds that have a value of 1 by default. Do not adjust Thresholds with default value of 5.**

Please refer to the BeadArray Controls Reporter Software Guide for more information on the calculations conducted to arrive at the output file results. This is an excellent documentation on calculations and interpretation of Methylation Controls.

How to prepare a Sample Sheet

• Be sure to reformat the barcode/Sentrix ID as a 'Number' with 0 decimal places in Excel before saving. Excel will default back to scientific notation, but the values should be in Number format before saving the .csv file.

Resources:

• BeadArray Controls Reporter Installer

• Illumina Knowledge: Sample Sheet Guidelines for GenomeStudio

For any feedback or questions regarding this article (Illumina Knowledge Article #3398), contact Illumina Technical Support [email protected].