Thank you for uploading that zip file.
Can you also send us the file "20230725_IntradaOA_Koley_90ACN_VR_TM_MC-TM-45_fullscan_neg_113_WT_con1.raw"?
I see that most of your chromatograms are zero most of the time and only have a non zero value at sporadic places.
When Skyline extracts chromatograms from spectra like this, Skyline sums up all of the intensities across a m/z window centered around your molecule's m/z.
The width of the channel that Skyline sums across is controlled by the "Resolving Power" setting at "Settings > Transition Settings > Full Scan".
I see that you have the resolving power set to 120,000. It might be that if you change that to a smaller number you will get better chromatograms.
After you change the resolving power you should tell Skyline to extract chromatograms again by pushing the "Reimport" button at "Edit > Manage Results".
By the way, if you want to see how wide of a channel Skyline is summing across, you can click on a point along the chromatogram. When you click on a point along the chromatogram, Skyline brings up a spectrum viewer which will show shaded regions around the places where Skyline summed intensities to extract chromatograms for particular transitions.
Also, for Thermo data, we usually suggest that you set the "Mass Analyzer" at "Settings > Transition Settings > Full Scan" to "Centroided" because Thermo's centroiding algorithm does such a good job of separating signals from things with neighboring m/z values. When the mass analyzer is "Centroided", the other setting becomes "Mass Accuracy" instead of "Resolving Power", but it still controls the width of the channel that Skyline sums across when extracting chromatograms.
-- Nick |