Basic Principles

 

Mass spectrometry was first successfully introduced in the 1940s for purification and accumulation of nuclear isotopes, and for analysis of volatile and low-molecular weight substances i.e. from petroleum (Polednak & Frome, 1981). In 1946, the concept of the time-of-flight technique was introduced by Stephens (Stephens, 1946) and the world´s first commercially produced mass spectrometer became available in 1948. It was based on the electronic ionisation (EI) technique and had a limited resolution with a maximum mass range of 300 Dalton. In the 1950s quadrupole techniques and the coupling of gas chromatography with mass spectrometry (GC-MS) were developed (Gohlke, 1959). At this time it was possible to analyse substances with a maximum mass of 1-2 kDa. In the late 1980´s, Tanaka, Karas and Hillenkamp were able to identify and define the masses and structures of the first larger biomolecules, by inventing a method that combined matrices with the laser-desorption ionisation technique (MALDI) and with time of flight mass analysers (TOF) leading to the MALDI-TOF technique (Tanaka et al. 1988; Karas & Hillenkamp, 1988).

 

MS in proteomics

The most commonly used tools for proteome analysis are 2D-PAGE followed by mass spectrometry via MALDI-TOF for protein identification via peptide mass fingerprint (PMF). Protein degradation by residue-specific cleavage into peptides is performed mainly enzymatically. The peptide masses can be acquired and afterwards matched against theoretical peptide libraries generated from the proteins as deposited in protein sequence databases. Due to the increasing demand for high-throughput proteome analysis, automated mass analysis and protein identification tools are already available (Gorg et al. 2000; Chamrad et al. 2003). Proteome analysis chips that utilise surface-coatings to enrich certain protein species from a sample are now available. When used in combination with MS (SELDI-TOF)  these appear to be an alternative to classical 2D-PAGE, but the chips currently lack reproducibility and are far less well validated than DNA-chips for example (Phelan & Nock, 2003; Jain, 2004).

 

Mass spectrometry in general utilises the differences in the mass-to-charge ratio (m/z) of ionised compounds or molecules for their separation in a high vacuum. The ionisation of the analyte is obtained through absorption or release of an electron. The ability to identify an unknown compound, determine its structure, and define its chemical characteristics in quantities as low as a pictogram, makes MS a universal tool. For the analysis of heterogenous mixtures, the mass spectrometer can be combined as a detector with gas chromatography (GC/MS) or liquid chromatography (LC/MS).

 

The main components of a mass spectrometer are an ion-source which generates the ions, a mass-analyser where these ions are separated, and a detector delivering the mass spectrum. After the application of the various existing ionisation and desorption methods used upon the molecules or peptides (ions, atoms or photons), generated ions will be dissociated in the vacuum of the mass analyser based on their m/z value.