Peptide shows two different peaks with in 2 min RT

Peptide shows two different peaks with in 2 min RT rajirathore3  2018-12-03

Hello skyline team

I am facing an issue with one of peptide which shows 2 different peaks for same peptide and contains same pattern of fragments. Data has been acquired on Thermo Q-exactive. This peptide elutes with in 2 min of RT and in some of the replicates it shows almost similar intensities and ppm error for both the peaks. This is a glucose modified peptid . Unmodified peptide is also acquired and that does not show this kind of pattern. I have attached spectra of few of the replicates which show only one peaks and other which show 2 peaks with almost similar intensities. I have attached a graph of retention time of replicate comparison. Please help me to resolve the issue

Brendan MacLean responded:  2018-12-03

We have seen peptides like this. There are actually 2 in the "Rat Heart Failure" dataset I like to teach at courses. I usually tell participants that these peptides would be extremely challenging to quantify without a heavy labeled peptide standard, and even then it is probably better to think of other peptides you might measure to get at the biology that is interesting you. If you absolutely need to measure this peptide form, then you should order an isotope labeled internal standard that will co-elute with it and you can use the ratio between the unlabeled and labeled forms as your proxy for quantity.

I wouldn't recommend using this peptide in a label-free experiment.