|Targeted MS/MS importing - cutting off ID'd peaks||kvancott2||2019-02-12|
I successfully built a spectral library from DDA data and then imported a list of proteins for an MRM-HR method using a Sciex 6600 QTOF.
I imported another DDA data file into my Skyline file. In the Transition Settings, I used "include all matching scans" option for Retention Time FIltering.
After importing the data file, the scan extraction width seems to be all over the place. Some peptides it is only a minute wide, some are 10 minutes wide, some are 20 minutes, and some even higher.
What is most frustrating, though, is that quite often the matching/ID'd peaks are cut-off in Skyline. I'm attaching a powerpoint file that shows an example of Skyline cutting off the scan of a yeast ADH peptide (YYVDTSK) that was spiked into the samples.
I've tried re-setting the Retention Time filtering, re-saving the file, re-importing the raw data file, but it doesn't change. I've tried adjusting the Retention Time filtering to 5, 10, 15, or 20 minutes - it doesn't seem to make any difference what the setting is.
I even tried importing one of the raw data files that was used to make the spectral library - same thing. Multiple peptides that are ID'd are cut off by Skyline. In your Targeted MSMS tutorial, it says on page 31 that the entire gradient should be covered in the chromatograms. But I'm not seeing that. Not sure what I'm missing.