Chromatogram information unavailable

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Chromatogram information unavailable yaoyao wang  2024-12-04 18:15
 

About 8 peptides were tested by waters QTof under MSMS mode. Peptide 1 resulted in "Chromatogram information unavailable" When I used skyline to analyze raw data, while others could work under same settings.

I am confused and want to know how to solve this.

I uploaded two peptieds information and results(peptide 1 and 2) in the file.

 
 
Nick Shulman responded:  2024-12-04 19:28
In Skyline you can use the menu item:
File > Share
to create a file whose name ends in ".sky.zip".
The .sky.zip file will contain the Skyline document that you attached as well as other important files including extracted chromatograms.

The "Share Document" dialog also gives you the option of including the raw files in the .zip file.

Please send us that .zip file as well as one or more of your .raw files.

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

In a PRM experiment, the usual reason that you would see "Chromatogram Information Unavailable" is that there really were no MS2 spectra where the isolation window matched the precursor m/z of the items in your Skyline document.
After we see your .sky.zip file and some of your .raw files we will probably be able to tell you what is going wrong.
-- Nick
 
yaoyao wang responded:  2024-12-04 20:46
Thanks for your timely reply. The zip file is attached as below.
 
Nick Shulman responded:  2024-12-05 03:40
Can you also send us one or more of your .mzML files?
That would be "pep2_MSMS_BEH_100ng [1A,3] [2].mzml" and/or "pep1_MSMS_BEH_100ng [1A,2] [1].mzml".

Those files will probably be too large to attach to this support request so you can upload them here:
https://skyline.ms/files.url
-- Nick
 
yaoyao wang responded:  2024-12-09 23:44
Excuse me for late response.

mzML files of two peptides were attached.
 
Nick Shulman responded:  2024-12-10 09:02
Thank you attaching those two .mzml files.
This appears to have been a DDA experiment where the mass spectrometer might have been told to fragment whatever m/z's it saw.
In the first .mzml file, spectra have been collected for precursor m/z's around 1132 and 1140.
In the second .mzml file spectra have been collected for precursor m/z's around 837.4.

I see that the Acquisition Method at "Settings > Transition Settings > Full Scan" is set to "PRM".
When the Acquisition Method is PRM, Skyline will extract a chromatogram point from MS2 spectra where the isolated m/z is within the "Method match tolerance m/z" of the m/z of the precursor in the document.
You can change the "Method match tolerance m/z" value on the "Instrument" tab at "Settings > Transition Settings".
Currently, that value is the default 0.055, but you could change that a value as high as 0.6.
After you change the Method match tolerance m/z to a higher number you could tell Skyline to extract chromatograms again using the "Reimport" button at "Edit > Manage Results".

If you tell us more about the sort of experiment that you were hoping to do we might be able to point you in a better direction.
-- Nick