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. Author manuscript; available in PMC: 2009 Aug 18.
Published in final edited form as: Nat Protoc. 2008;3(10):1630–1638. doi: 10.1038/nprot.2008.150

Figure 1. Diagrams illustrating the protocol.

Figure 1

(a) Scheme of the procedure for phosphopeptide enrichment and analysis. Protein extract is in-solution reduced, alkylated and digested with trypsin. Peptides are desalted and separated by SCX chromatography. Twelve fractions are collected, desalted, enriched by IMAC and analyzed by LC-MS/MS techniques. (b) Illustration of the StageTips used for combined IMAC enrichment and phosphopeptide desalting, where the IMAC resin, after incubation with peptide mixtures is finished, is packed on the top of C18 Empore disks. In the absence of ACN, phosphopeptides eluting from the IMAC resin are bound to the C18 disks, and salts can be removed before organic elution.