Hi Hossein,
This indicates that Skyline did not find any of the specified iRT standard peptides in your MaxQuant search results. Is it possible that you selected the wrong standards? Also, could you check the MaxQuant msms.txt file and verify that those peptides appear there (ctrl+f and search for one of the iRT standard peptide sequences)?

Thanks,
Kaipo

Hi Kaipo,
I checked for the iRT peptides and all 11 peptides are in my msms.txt. I can send you the files if you are interested.
Thanks
Hossein

Hi Hossein,
I was able to add the iRT peptides successfully. It looks like this is Pierce iRT standard peptides, is that right?
This is what I did:

1. Settings > Peptide Settings... > Library > Build
2. Choose where to save the library, select "Pierce (iRT-C18)" as iRT standard peptides
3. Add the input msms.txt file

Does that sound like the steps you took? If not, can you describe the steps you took to get to the error message?

Thanks,
Kaipo

Thank you very much for taking time and looking into this issue.
We didn't add Pierce iRT peptide mix to our samples and as you can see in the msms.txt we have Biognosys iRT peptides.
I have to mention that these samples are from yeast and the pierce peptides seem to be indigenous peptides that are part of the yeast proteome profile. It is very strange that skyline does see the pierce iRT peptides and not the Biognosys ones...
Any insights is greatly appreciated.
Hossein

Hi Hossein,
I couldn't find any Biognosys iRT peptides in the msms.txt file that you provided to me:
LGGNEQVTR
GAGSSEPVTGLDAK
VEATFGVDESNAK
YILAGVENSK
TPVISGGPYEYR
TPVITGAPYEYR
DGLDAASYYAPVR
ADVTPADFSEWSK
GTFIIDPGGVIR
GTFIIDPAAVIR
LFLQFGAQGSPFLK

But I found 13 out of the 14 Pierce iRT peptides in the file. Is it possible that you used the wrong msms.txt file?

Thanks,
Kaipo

Thank you so much Kaipo; this was really helpful. After re-visiting the Biognosys iRT peptides' sequence, it is now obvious that my version has AA shifts/mismatches. I am sure it will work fine after fixing the fasta file.
Hossein