SILAC-based quantitative proteomics (SILAQ) is an innovative technology used in high throughput quantitative analysis of large protein complexes, protein-protein and protein-small molecule interactions. SILAC service of Creative Proteomics provides an unbiased strategy that can reveal how specifically either inhibitors, or other perturbations, affect the dynamic properties and cellular distributions of proteins. It can also be used as a sensitive and effective method to determine the specific interaction partners of proteins in the cell.
Quantitative proteomics has increasingly gained impact in life science research as a tool to describe changes in protein expression between different cellular states. Stable isotope labeling by amino acids in cell culture (SILAC) is a powerful technique for relative quantification of proteins. Stable isotope labeling by amino acids in cell culture (SILAC) is a simple and straightforward approach for in vivo incorporation of a label into proteins for mass spectrometry (MS)-based quantitative proteomics. SILAC relies on metabolic incorporation of a given 'light' or 'heavy' form of the amino acid into the proteins. As the two isotopically labeled amino acids are essentially chemically identical, their incorporation does not interfere with normal cell growth, while leading to proteins/peptides that are distinguishable by mass and thus are ideal for mass spectrometric analysis. The method relies on the incorporation of amino acids with substituted stable isotopic nuclei. Thus in an experiment, two cell populations are grown in culture media that are identical except that one of them contains a 'light' and the other a 'heavy' form of a particular amino acid (e.g. 12C and 13C labeled L-lysine, respectively). When the labeled analog of an amino acid is supplied to cells in culture instead of the natural amino acid, it is incorporated into all newly synthesized proteins. After a number of cell divisions, each instance of this particular amino acid will be replaced by its isotope labeled analog. Since there is hardly any chemical difference between the labeled amino acid and the natural amino acid isotopes, the cells behave exactly like the control cell population grown in the presence of normal amino acid.
In comparation with derivatization-based labeling techniques directly after harvesting them for subsequent purification steps and analysis, the advantage of SILAC is that it ensures maximum reproducibility and minimum sample variation with regard to the protein level.
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