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Trying to figure out Skyline to put different concentrations of heavy peptide in calibration and in sample

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Trying to figure out Skyline to put different concentrations of heavy peptide in calibration and in sample divya-rathi  2025-02-26 03:15
 

Couple of queries:

  1. A colleague spiked 25 fmols of the heavy peptides in the standard curve but spiked in 20 fmols in the sample and I couldn’t find a place to have different values. I only saw one place to put in the concentration of the heavy peptides.

  2. A former colleague had 7 heavy and 2 regular synthetic peptides used for the calibration. When I look at the samples, I see the 2 regular synthetic peptides, but I am not finding any of the regular peptides that correspond to the remaining 5 heavy synthetic peptides and I am wondering if it was because they aren’t there or because they weren’t in the samples that I used for the calibration curve.

*We are new to Skyline.

Thank you for the help.
 
 
Nick Shulman responded:  2025-02-26 08:43
I am not sure I understand your first question.
For complicated dilution series you might need to provide "Explicit Analyte Concentration" values.
There is some information about that here:
https://skyline.ms/announcements/home/support/thread.view?rowId=63477

For your second question it might help if you sent us your Skyline document.
In Skyline you can use the menu item:
File > Share
to create a .zip file containing the Skyline document and supporting files including extracted chromatograms.
Files which are less than 50MB can be attached to these support requests. You can always upload larger files here:
https://skyline.ms/files.url

It also might help if you could provide a screenshot of what you were looking at.
--Nick