1. Introduction Mass spectrometry (MS) is an essential core technology in proteomics, the comprehensive analysis of the proteins expressed in a cell or tissue (Aebersold and Goodlett, 2001). In the late 1980s, two new ionization methods were developed that have revolutionized the analysis of large biomolecules by MS. Both methods, electrospray ionization (ESI, Fenn etal., […]

Core methodologies (Proteomics)

There is no science without fancy, and no art without facts – Vladimir Nabokov These are exciting times for proteomics. While technological progress at the frontiers of the science continues unabated, the basic ability to identify proteins in a mass spectrometer justifies much of the fuss. This section describes techniques that form the “bread and […]

Sample preparation for proteomics

1. Introduction Sample preparation can be defined as the process that goes from the biological sample of interest to the final protein extract that can be handled by the desired proteomics method. This process has to achieve parallel goals: • disruption of the initial structure of the biological material; • prevention of any spurious degradation […]

Tandem mass spectrometry database searching (Proteomics)

1. Introduction Mass spectrometry (MS) has become a principal technology in proteomics analysis largely due to improvements in sequence database availability and development of software tools to interpret mass spectra. Software tools of the modern proteomic era were first published around 1990 (Johnson and Biemann, 1989; Bartels, 1990; Yates et al., 1991). These programs facilitate […]

Interpreting tandem mass spectra of peptides (Proteomics)

1. Introduction Given the success of database search programs at identifying proteins (Clauser et al., 1999; Eng et al., 1994; Fenyo et al., 1998; Mann and Wilm, 1994; Perkins et al., 1999), one could reasonably question the importance of knowing how to manually interpret tandem mass spectra (MS/MS) of peptides. This is especially true when […]

FT-ICR (Proteomics)

1. Introduction It is Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (Amster, 1996; Comisarow and Marshall, 1974a,b,c; Jacoby et al., 1992; Marshall and Schweikhard, 1992; Marshall etal., 1998) which among mass spectrometric methods (see Article 10, Hybrid MS, Volume 5, Article 7, Time-of-flight mass spectrometry, Volume 5, Article 8, Quadrupole mass analyzers: theoretical and […]

Laser-based microdissection approaches and applications (Proteomics)

1. Introduction There is considerable heterogeneity in terms of both the numbers and types of cells present in different organs or tissues, reflecting their different biological functions. As an example, the cellular composition of the kidney comprises many diverse cell types including proximal tubule epithelial cells, distal tubule epithelial cells, collecting duct epithelial cells, endothelial […]

Orbitrap mass analyzer (Proteomics)

1. Introduction The orbitrap mass analyzer is rightfully considered to be one of the newest analyzers. Its roots, however, can be traced back to 1923, when the principle of orbital trapping was proposed by Kingdon (1923). Experiments over the next half a century have shown that charged particles could be indeed trapped in electrostatic fields […]

Time-of-flight mass spectrometry (Proteomics)

  1. Introduction Time-of-flight (TOF) mass spectrometers were introduced in the 1940s (Stephens, 1946; Cameron and Eggers, 1948) and commercialized in 1955 (Wiley and McLaren, 1955). Regarded for many years as instruments with low mass range and low resolving power, TOF mass spectrometers were “rediscovered” following the introduction of ionization techniques capable of ionizing biological […]

Quadrupole mass analyzers: theoretical and practical considerations (Proteomics)

1. Introduction Quadrupole mass analyzers are dynamic mass Alters that play a central role in mass spectrometric proteomic investigations. Quadrupoles are used both alone and in tandem for the isolation (by mass-to-charge) of ions; they may also be used (in RF-only mode) to contain ions during collision-induced dissociation (CID) (Yost and Enke, 1979). Gas-phase ions […]