Hi,

I am a farily new user of Skyline, and I am struggling to build my spectral library. Aside from some of the tutorials, my training is very limited. The source of my spectral library is a search from Proteome Discoverer 2.4 that I exported to a pep.xml file. I also have a used MSconvert to generate mzml files from my raw files.

I suspect that my issue is not due to any of the settings I made when setting up skyline, therefore I have not included those in this request. As seen in the attachment, I get an error saying the wrong directory is used, but I cannot seem to find out how it chooses this directory, or how to change it. I have put the skyline file, the raw files, and the converted spectrum files all in the same folder. The only thing missing, as I see it, is the program itself. However, I struggle to see the logic if this is the missing link, as this would be impractical for further projects.

I used the 20.2 version of Skyline and Windows 10 Home. The data I have is a non-targeted HPLC-MS/MS of CSF. I wish to use this library in a PRM project which is ready to be analyzed.

Thanks in advance! Let me know if I should add more info.
Ingrid