skyline shows PROSIT library twice in the mirrored image ignoring real data

skyline shows PROSIT library twice in the mirrored image ignoring real data pavel shliaha  2023-02-22 09:45

I am using a prosit library and I would like to see how my spectrum compares to the one in the library (predicted by skyline) using mirror image display. However smth is clearly wrong:

  1. in the mirror image the dotp 1, but in the target tab dotp is 0.89
  2. the name "2022_08_24_BSA_ATG4 vs prosit", but the 2022_08_24_BSA_ATG4 is the name of the library generated using prosit
  3. The mirror image includes the ions that I have removed from the analysis
  4. all the errors are 0 ppm implying that skyline just plot the library vs library

(see the screenshot attached).

Why is skyline not showing the actual spectrum from the current analysis as part of the mirror?

Nick Shulman responded:  2023-02-23 06:03
The "mirror" option in the Library Match window allows you to compare a spectrum in your spectral library (.blib file) with a spectrum which Skyline gets by asking the Prosit web server for a spectrum.
It sounds like your spectral library .blib file was generated from Prosit, so, yes, you would expect the top and bottom halves of your mirror spectrum to be identical.

It sounds like you would like Skyline to instead be showing you a spectrum from your raw file. I cannot think of a way to get Skyline to do that for you.
If you want to see how your chromatogram peak areas compare to the intensities in your spectral library, you can use the menu item:
View > Peak Areas > Replicate Comparison
The peak area replicate comparison graph will show you one bar representing the intensities from the spectral library, and it will also have other bars for the replicates in your Skyline document.
-- Nick
pavel shliaha responded:  2023-02-24 06:03
thank you! It would be great to have a similar mirror view for library vs real data. It is hard to judge fragment intensity agreement from those barplots