Stefanie,

I can't figure out how to get the chromatograms in your Skyline document from the .mzXML files that you uploaded.
The Skyline document is here:
https://skyline.ms/_webdav/home/support/file%20sharing/%40files/SIM_Analysis.sky.zip
and the .mzXML files are here:
https://skyline.ms/_webdav/home/support/file%20sharing/%40files/GCMS_Sim.zip

When I tell Skyline to extract chromatograms (File > Import > Results) from the .mzXML files, all I see are completely flat chromatograms.

Are the settings in the Skyline document different from what they were when you originally told Skyline to extract chromatograms? Or are the .mzXML files different from the files that the chromatograms were extracted from?

When I look at those .mzXML files using ProteoWizard SeeMS.exe, they seem to all be MS1 spectra, and I am not sure how Skyline might have been interpreting them to give you the extracted ion chromatograms that you have.
-- Nick

I later added files to the folder I sent to you, but the document is the same and the setting have not been changed. The updated file-folder is named "SIM-Messungen_Wiederholung_2". When I import the folder with the added files into skyline, nothing is found. Before at least some fragments were found (see attached screenshot).

I will try again later with the updated version of skyline. If you have some further input, I'd be grateful.

- Stefanie

Stefanie,

I just noticed that in the Skyline document you sent us, the "Triggered chromatogram extraction" checkbox has been checked on the Instrument tab at "Settings > Transition Settings".
You should uncheck that checkbox.
That checkbox is only supposed to be used with things like Thermo's SureQuant methods where the mass spectrometer has been told to only acquire data for some targets after other things have happened.
When the checkbox has been checked, Skyline tries to detect the time ranges over which data was not being collected.
It completely messes things up if you have that checked and it was not a SureQuant method.

After you have unchecked that checkbox you will need to tell Skyline to extract chromatograms using perhaps the "Reimport" button at "Edit > Manage Results".

Let us know if you are still having trouble with your dataset after unchecking the "Triggered chromatogram extraction" checkbox.
-- Nick

Uncheck the "Triggered chromatogram extraction" helped a lot. I have at least one fragment per molecule (see attachment). For now that is enough to go on.

Thanks a lot for your support!
Stefanie

Stefanie,

The chromatograms that Skyline extracted are very spiky, but the default chromatogram display option in Skyline hides that from you.
If you right-click on the chromatogram graph and choose "Transform > None" you will see that the signal for each transition alternates between high values and low values.
If you click on a point along the chromatogram Skyline will show you the spectrum that contributed to that chromatogram point.
Each spectrum only has signal for the m/z that was isolated, which I guess is to be expected for SIM scans.
Unfortunately, Skyline does not recognize these as SIM scans. I believe the reason that Skyline does not recognize these as SIM scans is that the scan window width is zero (e.g. "109-109") which Skyline seems to be interpreting as no information.

How exactly did you create these .mzXML files? Could the tool that you used to create the .mzXML files have created .mzML files instead?
.mzML files can contain more information than .mzXML files, so it is possible that there would be a way to get your data to work with Skyline if you had .mzML files.
If you are able to create .mzML files, please send them to me and I will see whether there is a way to get the .mzML files to work with Skyline.
-- Nick

Dear Nick,

I used MSConvert by ProteoWizard to convert the .mzXML files to .mzML files and uploaded a zip-folder to the file-section: "Pesticides_SIM_all.zip".

Would be great if that works better. Thanks again for trying to find a way to get my data work!
Stefanie