Get hands-on experience creating a Skyline document to measure quantitative differences in peptide expression using the MS1 scans from your data dependent acquisition (DDA) experiments. In this tutorial, you will generate a spectral library from a discovery data set, set up a Skyline document for MS1 filtering, import raw mass spectrometer data to extract precursor ion chromatograms from MS1 scans, with peak picking guided by MS/MS peptide identifications, and further process the resulting quantitative data in Skyline. If you are interested in label-free quantitative analysis of discovery data sets, this tutorial will give you a new tool set for your investigation. (41 pages)
* - written on v1.2, updated for v1.4, revised for v2.5, updated for v20.1, v21.1, and v22.2
Also, see our paper in Molecular Cellular Proteomics (please cite)
Platform independent and label-free quantitation of proteomic data using MS1 extracted ion chromatograms in skyline. Application to protein acetylation and phosphorylation
On October 21st, 2014, the Skyline Team produced Webinar #1: Getting the Most Out of DDA Data with Skyline, another great resource for working with DDA data in Skyline.
On June 16th, 2015, the Skyline Team produced Webinar #8: DDA to Targeted: Differential Statistics with Skyline, An untargeted approach starting from all peptides detected in a DDA experiment reduced to proteins that appear to be changing in the samples.
On September 29, 2015 the Skyline Team produced Webinar #10: Working with Modifications in Skyline , with an in-depth discussion peptide modifications, importing PTMs, isotope labeling and importing large assay libraries, among other topics.
If you run into trouble seeing ID annotations in your chromatograms, be sure to consult Tip: ID Annotations Missing with Mascot Search Results. Even if you did not use Mascot, this tip may contain useful information.