MS1 filtering: is manual XIC window mass tolerances possible?

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MS1 filtering: is manual XIC window mass tolerances possible? alejandro.cohen  2020-10-23
 

Hi Skyline,

I'm using Skyline for a targeted metabolomics project on an QExactive (running at 70K resolution @ 200m/z according to Thermo). I inserted this value in the Transition Settings> Full-Scan>MS1- filtering>Resolving Power tab.

My standards are all showing at the expected Rt and with mass errors <2ppm. However, Skyline is also labeling other peaks between + - 2 to 9ppms. Is there any way to manually reduce the XIC mass windows, or set mass error thresholds to reduce the 'labeling clutter' in the chromatogram windows? Or do I just have to increase/fake the Resolving Power value on the Transition Settings pane.

Thanks again for you continuous support, Skyline rocks!

Alejandro

 
 
Brian Pratt responded:  2020-10-23

Hi Alejandro,

If you are confident in your chromatography you can set explicit RT and RT windows for each precursor in your imported transition list.

But yes, you may wish to adjust Skyline's resolving power first and see if that works for you - it's certainly the more convenient approach.

Thanks for using the Skyline support board,

Brian Pratt

 
alejandro.cohen responded:  2020-10-23

Thanks Brian for you quick response.

Would this be a good suggestion for developers to add to skyline: -User defined XIC mass windows?

Although you can add the Resolving power of instrument, it is unclear how Skyline 'calculates' this window value (or at least now what that window is would be a good start). Having to 'tamper' the Resolving Power to remove those other peaks seems a little awkward and requires a 'test-and-see' which is time consuming.

The Explicit RT would work, albeit some risk in chromatgraphic drift.

Your thoughts?

Alex

 
Brian Pratt responded:  2020-10-23

Hi Alex,

Perhaps you could supply a screen shot of the issue you're experiencing?

The Transition Settings > Full Scan > MS1> Resolving power isn't an instrument setting, it's a chromatogram extraction filter setting that you can use to tune the automatic peak integration. It needn't match the machine setting, though that's probably a good place to start.

Can you elaborate on what you're thinking of for your new feature idea?

Also, have you had a look at the "Use high selectivity extraction" setting in the Transition Settings > Full Scan window?

Thanks

Brian

 
alejandro.cohen responded:  2020-10-23

Brian,

-"The Transition Settings > Full Scan > MS1> Resolving power isn't an instrument setting, it's a chromatogram extraction filter setting that you can use to tune the automatic peak integration. It needn't match the machine setting, though that's probably a good place to start."

  • I am aware of this. However, this should 'ideally' match your acquisition parameters I guess. It would be nice to know what is the XIC mass window (in ppms or Da) Skyline is using for the values used in this setting. Or alternatively, have a 'Manual Extraction Filter' where the user can choose/fix the mass window for the integration instead of entering a Resolving Power... I think I'm not very clear, or more likely, not understanding what boundaries skyline is using to integrate the peaks.

-"Also, have you had a look at the "Use high selectivity extraction" setting in the Transition Settings > Full Scan window?"
-Yes I have, and it is checked.

Regardless, I think we can close this thread now.... it seems that when initially I changed the MS Resolving power as you suggested, the changes were not applied directly at that moment (even after hitting Reintegrate option). I now just recently closed and reopened the files and those other background/baseline chromatographic peaks I mentioned earlier have disappeared :)

Nice weekend!

 
alejandro.cohen responded:  2020-10-23

Brian,

I think I have the answer from Brendan's reply in the thread entitled: "Precision of MS1-XICs" thread.

Skyline's algorithm is more complex than just integrating a fixed m/z window around a theoretical m/z value. I guess I underestimated the complexities under Skyline's hood.

Cheers

Alex