can i use skyline to process product ion MS2 data to select one product ion to do quantitation

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can i use skyline to process product ion MS2 data to select one product ion to do quantitation xue shi  2022-07-06
 

Dear customer service,
I am running product ion(MS2) experiment on thermo orbitrap. I want to process data with skyline. However, I can only process the SIM and MRM data but don't know how to process MS2 data for quantitation. Could i view spectrum by skyline and pick the strongest product ion to quantitation with skyline? Is there any video or manual can help with this?
Thank you a lot.
Xue(Katie) Shi

 
 
Nick Shulman responded:  2022-07-06
If you want to do MS2 quantification, your acquisition method should either be PRM or DIA.
In a PRM experiment, you give the mass spectrometer a list of precursor m/z's and the mass spectrometer collects many MS2 spectra with those precursors.
Here is our PRM tutorial:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_prm
It sounds like you already know which molecules you are interested in collecting data for, so PRM is probably what you want to do.

In a DIA experiment, you tell the mass spectrometer to isolate wide windows which together cover a wide m/z range.
We have several DIA tutorials. This is probably the best one to start with:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_dia
If you did not know which molecules you wanted to monitor, but instead wanted to collect data for everything, DIA would be the right choice.
-- Nick
 
Brendan MacLean responded:  2022-07-07
The answer to the second question: "Could I view spectrum by skyline and pick the strongest product ion to quantitation with skyline?" is Yes.

Skyline provides two possible ranking values in its reports:

Transitions.Library Rank - ranking based on intensity of a spectrum in a spectral library
TransitionResults.Peak Rank - ranking based on imported data

You should consult the tutorial on the Skyline Live Reports to understand how to use them with the Document Grid:

https://skyline.ms/tutorial_custom_reports.url

The other important property for what you mention is:

Transitions.Quantitative

Once you understand these and how to use the Document Grid, it is relatively straightforward to create a report that includes them, filter by a ranking and turn off the Transitions.Quantitative property for all transitions with the rankings you do not want included in your quantification.

Thanks for your interest in Skyline.

--Brendan