CPC Scientific

Isotope Labeled Peptides

Genomics research shows that more than a million proteins are encoded by approximately 30,000 human genes. Proteomics, the study of proteins encoded by the genome, includes identification of post-translational modifications, structural analyses, protein localization studies, and protein quantitation. Mass spectroscopy-based techniques have evolved as a powerful tool in proteomics. Stable isotope-labeled peptides (SIL peptides) are chemically and physically indistinguishable from their endogenous counterparts with respect to retention time, ionization efficiency, and fragmentation pathways. Therefore, either are ideal internal standards and template analytes. Peptides can be labeled with one or more isotopes of hydrogen, carbon, nitrogen, or oxygen by incorporating amino acids containing the desired isotopes such as deuterium (D), 13C, 15N, or 18O into the peptide during synthesis.

The Absolute Quantification method (AQUA) enables targeted quantification of protein and post-translational modifications in complex protein mixtures using SIL peptides as internal standards. The SIL peptide is introduced into a biological sample during or after protease digestion. The heavy SIL peptide and its light endogenous peptide fragment are detected by selected reaction monitoring (SRM) in a mass spectrometer. Based on the known amount of SIL peptide added, and the intensity ratio of both peptides, the amount of endogenous protein can be calculated.