Product ions filtering in DIA

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Product ions filtering in DIA Kavee  2018-05-06 18:44
 

Hi,

I'm interesting in doing DIA experiment and have watched the skyline webinar 14,15. In the transition setting, you suggest to use product ions from '3' to 'last ion'. Could you explain the reason to exclude y1,y2 and b1,b2 ions? I try to find the answer for quite some time but still don't understand. Another question is could I include p ion in the ion types?

Thank you very much.
Kavee

 
 
Nick Shulman responded:  2018-05-07 11:04
Yes, including the precursor ion types is a good idea.

The reason that you would want to exclude the small ions is that they are likely not going to be specific to the peptide of interest. Specifically, if you used trypsin to digest your proteins, then nearly all of the y1 ions that you see will just be the amino acid "K" or "R". Therefore, the y1 ion that you see in a DIA MS2 scan might have come from the peptide that you are interested in, but it also could have come from any other peptide whose precursor m/z was within the width of that isolation window.
 
Brendan MacLean responded:  2018-05-07 11:39
Note that you can also exclude small ions by setting the Min m/z value on the Transition Settings - Instrument tab to something like 300 (or 250 to 350). I know the Aebersold lab does this in preparing libraries for OpenSWATH, and it may be a more direct way of excluding low m/z ions, since if you allow high enough fragment ion charge states, eventually even larger ions can become low m/z.
 
Kavee responded:  2018-05-07 13:31
Thank you very much.