While LC-MS/MS has been used for the identification of proteins from complexes and cell lysates (qualitative proteomics), until recently the quantitative study of gene expression using differential display has been restricted to 2-D gel analyses. However, an alternative non-gel based approach has been the use of isotope coded affinity tags (ICAT) for the quantitative study of gene expression at the proteome level. Recently, an improved approach analogous to ICAT has been developed called iTRAQ (Applied Biosystems)

The technique is based upon chemically tagging the N-terminus of peptides generated from protein digests that have been isolated from cells in, for example, two different states. The two labelled samples are then combined, fractionated by nanoLC and analysed by tandem mass spectrometry. Database searching of the fragmentation data of the peptides results in the identification of the labelled peptides and hence the corresponding proteins. Fragmentation of the tag attached to the peptides generates a low molecular mass reporter ion, that is unique to the tag used to label each of the digests. Measurement of the intensity of these reporter ions, enables relative quantification of the peptides in each digest and hence the proteins from where they originate.There are four tags available enabling four different conditions to be multiplexed together in one experiment. Please contact us for details.