Figure 1

Workflow of `bottom-up’ or shotgun proteomics. Protein extracts from cells, tissue or biofluids are prepared by mechanical (e.g., glass bead or homogenization) or chemical-based (precipitation, detergent solubilization) methods. Proteins are proteolytically digested into peptides, usually with trypsin, that are separated by 1D or 2D chromatographic separation. The final chromatographic step is performed in-line with the mass spectrometer. Two scan types are acquired: MS1 spectra contain intact peptide mass to charge (m/z) values; MS2 or tandem MS (MS/MS) spectra represent peptide fragment ion m/z values. Peptide MS1 and MS2 data are correlated with theoretical peptide m/z values with database search programs that use protein sequences as templates; parsimonious protein identifications with peptide matches are reported.