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Inquiry on peak data points, TIC normalization, and reporting in Skyline

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Inquiry on peak data points, TIC normalization, and reporting in Skyline jgt4047  2026-03-27 06:28
 

Dear Skyline Team,

I hope you are doing well.

I am currently performing PRM data analysis using Skyline and would like to ask a few questions regarding data extraction and reporting within a consistent quantitative workflow.

I would like to obtain the number of data points per peak within the user-defined retention time (i.e., within the peak integration boundaries) and include this information in a report.Could you please advise which fields or columns in the Document Grid should be used to extract this information?

I am planning to perform TIC normalization and would like to export the normalized values for reporting.Could you please let me know which fields should be added in the Document Grid to obtain normalized intensities?

In addition, I would like to directly inspect the peak area within the selected integration boundaries as well as the corresponding TIC values, in order to verify the normalization process.Could you please advise which fields in the Document Grid allow access to these values?

I would greatly appreciate your guidance.

Thank you very much for your support.

Best regards,
 
 
Nick Shulman responded:  2026-03-27 07:13
The column in the Document Grid which will tell you the number of data points between the integration boundaries is "Points Across Peak". That number is reported per Transition and Replicate since in an SRM experiment each Transition might have a different number of points per peak. In a PRM experiment, all MS2 transitions for a given precursor would be expected to have the same number of points across the peak.

Skyline can do the TIC normalization for you. If you go to the "Quantification" tab at "Settings > Peptide Settings", you can choose "Total Ion Current" as the "Normalization Method".
If you do that, then the "Normalized Area" value in the Document Grid will be the area normalized to the total ion current.
When Skyline normalizes to Total Ion Current, Skyline multiplies the measured values by the median Total Ion Current Area across all the Result Files, and divides by the Total Ion Current Area for that particular Result File.

If you want to do this calculation yourself you can add the "Total Ion Current Area" column to your report.
-- Nick