Detection Q Value shows #N/A for solid peptides

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Detection Q Value shows #N/A for solid peptides Z. Jiang  2021-02-05
 

Dear Skyline Team,

I am currently working on analyzing DIA data on Skyline. One thing I noticed is that quite a lot peptides with very good XICs and dotp scores have "Detection Q Value" shown as #N/A. For example, attached figure (Detection Q Value NA.png) shows the XICs of one of the iRT peptides. It is definitely in my sample and has very well-aligned XICs, intensities and dotp scores. However, the detection Q value is #N/A. Actually, all my iRT's detection Q values are #N/A. Does #N/A mean that the peptide is not significant?

I performed "Refine - Reintegrate" and trained "mPhrophet" model with target-decoy strategy (see figure "Skyline_Reintegrate.png"). After that, I selected "Only integrate significant q values" with a cutoff of 0.01. Did I do anything wrong?

Thank you,
Zhenze

 
 
Nick Shulman responded:  2021-02-05
The detection Q values of your iRT standards are supposed to be #N/A.

The standard peptides in your Skyline document (i.e. the QC standards, Global normalization standards and iRT standards) are expected to behave differently than the regular peptides in your samples, so Skyline excludes standard peptides from the whole process of training the model and calculating Q values.

When you tell Skyline to "only integrate significant q-values", the things that do not get integrated will still have their high Q-values.
The "Area" values will be "#N/A" for the things that did not get integrated.

If you have any other questions, you should send us your Skyline document so we can see what you're looking at.

In Skyline you can use the menu item:
File > Share
to create a .zip file containing your Skyline document and supporting files including extracted chromatograms.

If that .zip file is less than 50MB you can attach it to this support request.
Otherwise, you can upload it here:
https://skyline.ms/files.url

-- Nick