PRM Workflow with Spectral Matching

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PRM Workflow with Spectral Matching sa825  2019-10-18
 

Good afternoon,

I have watched Webinars 3 and 4 and found it very helpful but I have a question regarding spectral matching.
I conduct PRM on the Thermo Q Exacitive and this is my current workflow:

  • I choose the precursor and product ions of interest and export an isolation list for the Q Exacitive
  • I get the Raw Data file from the Thermo and run it through PD (with Percolator) which produces a PD result file
  • I then import this PD result file as a PRM search (File -> Import -> Peptide Search) and match it with the same PRM Raw file by following the instructions in the Import Wizard
  • This produces what I would call really good spectra. The chosen product ions are clearly detected with Peptide ID annotations.

However, when I import the raw result file ( File -> Import -> Results), I don't get the same level of detection. In most cases, none of the product ions detected when I do the workflow above are seen in this result file approach.

Can you please explain why there is this discrepancy? And if my current workflow is correct?

Thanks,

Shimon

 
 
Nick Shulman responded:  2019-10-18
Can you send us your Skyline document and that problematic .raw file?

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.

You can upload that .zip file and your .raw file(s) here:
https://skyline.ms/files.url
-- Nick