Figure 19. Overview of the integrated digestion and labeling procedure on immobilized trypsin beads. The sample is first digested in ammonium hydrogen carbonate buffer (in H216O) followed by evaporation. The sample is then reconstituted in H218O containing buffer (pH 6) and extra trypsin beads (separate vial). Labeling is then carried out in the same vial as the digestion, and is stopped by removing trypsin beads and adding 8 M urea. Corresponding 16O- and 18O-labeled samples are mixed in a 1:1 ratio before LC-MS/MS analysis
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