Mass spectrometry (MS) has flourished over the last 10-15 years with the advent of "soft" ionization techniques to measure the molecular masses of proteins and peptides. The principal techniques used are electrospray ionization mass spectrometry (ESI-MS) and matrix-assisted laser-desorption ionization mass spectrometry (MALDI-MS).

The main advantage of ESI-MS is the generation of ions in an organic solvent allowing the mass spectrometer to be coupled directly to a HPLC system for on-line analysis during peptide separation.
MALDI-MS has the advantage of being more tolerant to the presence of salts and detergents, allowing rapid, high-throughput analysis of digests of protein spots from 2-D gels. Both techniques allow the accurate mass measurement of intact proteins and the acquisition of sequence information from peptide fragmentation spectra for protein identification.

In conjunction with the growing number of genome sequences that are available in databases, MS can be used to investigate the proteomes of intact cells and subcellular fractions from peptide fragmentation data. This can be conveniently performed either by analyzing spots from 2-D gels by MALDI-MS-MS or by 2-D HPLC-MS-MS on ion-exchange and reverse-phase columns ("gel-free proteomics"). Moreover, information on changes in protein expression can be obtained by measuring changes in spot intensities in 2-D gels from treated and untreated cells (2D-DIGE technology) coupled to protein identification and by differential isotopic tagging or labelling of peptides prior to 2D-LC-MS.

Given the limited number of genes predicted in some species, functional diversity must be created through other means. Since genomes databases provide few clues as to the existence of splice variants of proteins and post-translational modifications (phosphorylation, glycosylation, acetylation, ubuquitylation...), one of the great challenges in proteomics in the post-genomic era is to use MS to pin-point these variations and hopefully quantify their changes.