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Mass spectrometry (MS) has become a vital enabling technology in the life sciences. This chapter summarizes the fundamental aspects of MS, with reference to topics such as isotopic abundance and accurate mass and resolution. A broad and comprehensive overview of the instrumentation, techniques, and methods required for the analysis of biomolecules is presented. Emphasis is placed on describing the soft ionization methods and separation techniques employed in current state-of-the-art mass spectrometers.

Membrane proteins constitute a substantial fraction of the human proteome. A small subgroup associates with membranes through the presence of a C-terminal lipid anchor that is joined to the protein via a phosphoglycan. The prion protein (PrP), an abnormally folded form that causes fatal neurodegeneration, is one example of a glycosylphosphatidylinositol (GPI)-anchored protein. Although GPI-anchored proteins were first recognized some 20 years ago (in the mid-1980s), relatively few GPI anchors have been analyzed in detail. Therefore, a description of the analysis of the PrP-GPI anchor using a variety of mass spectrometric methods is of interest even though some of the approaches adopted could be facilitated through the use of newer, more sensitive techniques.

The new technologies in mass spectrometric analysis of peptides and proteins necessary to accommodate proteomics-scale analyses require, in turn, concomitant development of informatics technologies suitable for very large-scale data handling and analysis. This chapter focuses on the data analysis tools available to the community for analysis of mass spectrometric proteomics data. Different database searching strategies are discussed for peptide and protein identification, and approaches and tools available for comparative quantitative analysis of samples are outlined.