Protein relative quantification

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Protein relative quantification tiziano brogna  2019-07-03
 

Hi,

I have 2 biological conditions for which we monitore multiple proteins to check if they are up or down-regulated.
I have analytical replicates and we monitore multiple peptides for each proteins.

I would like to know how Skyline consider the variability for the different peptides used for protein relative quantification (at the end we obtain fold change ratio with error bar that are linked with the analytical replicates if I understood well)?

I based the method on this tutorial:
https://skyline.ms/_webdav/home/software/Skyline/@files/tutorials/GroupedStudies1-3_1.pdf

In other word, I have the variability for the analytical replicates and now I would like to know how Skyline use the peptides signals to give the fold change information for the 2 conditions tested.

Thank you,

Best regards,

Tiziano

 
 
Brendan MacLean responded:  2019-07-06

Hi Tiziano,
Skyline uses pretty simple statistics. It sums peak areas for all peptides of a protein and averages technical replicates, dropping values with missing data. Then it performs a T-test on the "subjects".

For a more sophisticated statistical approach which treats each transition of the protein separately and uses actual imputation of missing values, we suggest you use the MSstats R implementation. You can start with the Skyline External Tool, which you can install with Tools > Tool Store, or you can download from here:

https://skyline.ms/skyts/home/software/Skyline/tools/details.view?name=MSstats

Good luck with your group comparisons.

--Brendan