Improved In-Gel Digestion Results and Work-Flow Through the Use of a Mass Spectrometry Compatible Surfactant
Part # PS094
Daniel J. Simpson1, Sergei Saveliev1, Becky Godat1, and Grzegorz Sabat2
1Promega Corporation, Madison, WI
2University of Wisconsin, Madison, WI
Typical protocols for in-gel protein digestion require overnight incubation followed by additional 2–3 hours for peptide extraction. We present here a protocol and reagent to streamline this process and in many cases improve protein coverage. The reagent, ProteaseMAX™ Surfactant, enhances peptide cleavage by providing a denaturing environment prior to protease addition. It is also designed to degrade during the proteolytic reaction, generating species that do not interfere with mass spectrometry. For in-gel applications, complete digestions can be performed in 1 hour and the surfactant provides concurrent peptide extraction. This obviates the requirement for separate extraction processing, greatly simplifying the overall work-flow. Recovery is particularly enhanced for longer and more hydrophobic peptides (2,500–4,000Da), thereby improving coverage.
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