MS1 filtering

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MS1 filtering paul.derbyshire  2021-01-28
 

Hi Skyline team,

I am having a go at MS1 filtering in Skyline and seems to be working quite well. If I click on the chromatogram peaks a new window opens up showing the precursor and it's isotopes (M, M+1, M+2 and so on). If I zoom in to say, the precursor (see attaches image file) I see that this peak is made up of other peaks. I am assuming these are subtle isotopic variants around this m/z and reflect their natural occurrence. Is this interpretation correct? Look forward to your reply.

Thanks

Paul

 
 
Nick Shulman responded:  2021-01-28

The vertical blue lines within that Gaussian shape indicate the exact m/z values that the mass spectrometer reported intensities for.

That is, a profile scan is represented as a list of m/z and intensity values. For instance, here is a small Gaussian shape around 425.23:

m/zIntensity
435.22171541904034
435.224001212509191
435.22628730561540
435.228572844725704
435.230858744618120
435.233144630828632
435.235430513942950
435.23771653564551

These are the only numbers that Skyline got from the mass spectrometer. You are supposed to interpret that as a continuous signal. That is, you are not supposed to think that the signal dropped to zero between 435.2217154 and 435.2240012.

Skyline calculates the peak area by summing all of the intensities for the m/z's which in a certain range around the calculated m/z of your peptide. The width of the channel that Skyline sums across is controlled by the resolution settings that you have at "Settings > Transition Settings > Full Scan".

Note that the Gaussian shape of this curve is a consequence of the way that mass spectrometers measure things. The vast majority of the ions that contributed to that shape had exactly the same mass to charge ratio, but everything was smeared out because of the way that the physics inside of the mass spectrometer works. There is a tiny amount of variation caused by molecules that have a heavy hydrogen instead of a heavy carbon, but these mass spectrometers are not precise enough to be able to see that.

Does that answer your question or were you asking something else?
-- Nick

 
paul.derbyshire responded:  2021-01-29

Hi Nick,

Thank you for the clear explanation which answered my question. I'll play around with the resolution settings to see what effect this has on channel width.

Cheers

Paul