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Chromatogram Information Unavailable olivia schwartz  2025-06-23 16:10
 

Hello!

I've been learning to use Skyline and wanted to, for now, test inputting a transition list with just the internal standard but keep getting "Chromatogram Information Unavailable" when uploading files that I've confirmed have the internal standard detected well. This was data from a Q Exactive HF using DDA, and the internal standard is DL-Tyrosine (3,3-Dâ‚‚) (HOC6H4CD2CH(NH2)COOH).

I also notice if I omit the precursor and product m/z columns, Skyline will denote 186 as the precursor instead of 184, so I am not sure if there is a discrepancy in my transition list?

I have attached an .mzML that should show detection for this compound along with the transition list I tried.
Thanks!
 
 
Nick Shulman responded:  2025-06-23 16:40
I see that you uploaded a .mzML file containing DDA data.
Skyline is not a good tool for looking at DDA data because everything Skyline does revolves around extracting chromatograms.

With DDA data, any particular precursor will only sporadically be selected for fragmentation, and so, if you do get a DDA MS2 chromatogram, it will have long straight lines connecting the points where a matching precursor was fragmented.

It is possible to extract MS2 chromatograms from DDA data using Skyline, but only if you have also told Skyline to extract MS1 chromatograms. In that case, the purpose of the MS2 chromatograms are to give you something to click on so that you can look at the MS2 spectra, but the real thing that Skyline wants to do is show you the MS1 chromatograms and their peak areas.

Skyline has a lot of tutorials to help you get started:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorials

If you are working with small molecules then I would recommend the small molecule targets tutorial:
https://skyline.ms/wiki/home/software/Skyline/page.view?name=tutorial_small_molecule

-- Nick