  Sequence coverage in MS analysis of protein digestion-derived peptides is a
key issue for detailed characterization of proteins or identification at low
quantities. In gel-based proteomics studies, the sequence coverage greatly
depends on the protein detection method. It is shown here that ammoniacal
silver detection methods offer improved sequence coverage over standard silver
nitrate methods, while keeping the high sensitivity of silver staining. With
the development of 2D-PAGE-based proteomics, another burden is placed on the
detection methods used for protein detection on 2-D-gels. Besides the classical
requirements of linearity, sensitivity, and homogeneity from one protein to
another, detection methods must now take into account another aspect, namely
their compatibility with MS. This compatibility is evidenced by two different
and complementary aspects, which are (i) the absence of adducts and artefactual
modifications on the peptides obtained after protease digestion of a protein
detected and digested in - gel, and (ii) the quantitative yield of peptides
recovered after digestion and analyzed by the mass spectrometer. While this
quantitative yield is not very important per se, it is however a crucial
parameter as it strongly influences the S/N of the mass spectrum and thus the
number of peptides that can be detected from a given protein input, especially
at low protein amounts. This influences in turn the sequence coverage and thus
the detail of the analysis provided by the mass spectrometer.
