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P roteomics is defined as the protein complement of the genome and involves the complete analysis of all the proteins in a given sample [1,2]. Several technologies are involved, and numerous questions concerning the proteins are addressed. What proteins are contained in a biological sample? At what concentration do the proteins exist? How do protein expression levels alter in different samples? What are the posttranslational modifications (PTMs)? Where in the cell [3] or an organism [4] are the proteins localised? How do the proteins interact with other proteins or molecules [5,6]? The following discussion concentrates on computational aspects of protein identification. Characterization (identification of protein modifications), quantitation, and sample comparisons are also discussed briefly. A typical proteomic experiment involves the analysis of complex samples, i.e., containing many proteins at varying concentrations [7]. Most of the currently available technology for identifying proteins from biological samples simply cannot contend with the complexity, and the majority of the low-abundance proteins are not observed. There are, however, a number of methods to separate the proteins contained in the original sample to obtain a simpler sample set that is amenable to in-depth analyses. Typical technologies are electrophoretic gels [8] and liquid chromatography [9] (LC) (see Figure 1A). A dominant and well-practiced technique in proteomics is referred to as the ‘‘bottom-up’’ approach. Proteins are digested into peptides (smaller components of the protein) by a proteolytic enzyme, e.g., trypsin. Analysis of the peptides is achieved by mass spectrometry (MS), and, from the data generated, the peptides (and subsequently the proteins) can be identified. The resultant mixture of peptides obtained from the digestion of several proteins is often highly complex, and a degree of separation can be achieved by peptide LC. Possible combinations of separation techniques are illustrated in Figure 1B. Mass spectrometers comprise three main components: an ion-source, a fragmentation cell, and a mass analyzer. Each component is essentially independent from the others, and as such it is possible to combine the different technological aspects to produce different types of mass spectrometers. To measure its molecular mass, a molecule must be ionised. This occurs in the ion source of the mass spectrometer. The source can be based either on electrospray ionization [10] (ESI), which is therefore appropriate for liquid samples; or on matrix assisted laser desorption ionization [11] (MALDI), which is appropriate for samples that have been mixed with a matrix and crystallized on a metallic plate. The most common types of mass analyzers used in proteomic laboratories are (i) ion trap (IT), where the radio frequency of the trap is varied and the ejected ions are detected; and (ii) time-of-flight (TOF) analyzers, where the time required for an ion to ‘‘fly’’ through an electric field–free region of the instrument is recorded and correlated to the mass of the ion. Most current instruments include a fragmentation cell that uses an inert gas to break the peptides by collision-induced dissociation (CID). A fragmentation cell, however, is not always present (see next section), or fragmentation can occur ‘‘spontaneously’’ (in-source and post-source decay). All mass spectrometers do not measure mass directly, but rather the mass-to-charge ratio. Hence the measurements obtained are dependent on the charge state(s) of the molecule.