Targeted Proteomics

Verification of potential biomarkers is done using Targeted Proteomics with Multiple Reaction Monitoring (MRM). The targeted MRM proteomics technology is depicted in Figure 1, and is based upon selecting at least two predicted tryptic peptides from each protein of interest and then synthesizing heavy and light stable isotope versions of these “standard” peptides. After synthesis, a standard curve is generated from precise quantities of each of these internal standard, synthetic peptides. The heavy synthetic peptides are then spiked into the sample at known concentrations. LC-MRM analysis is then performed on a triple quadrupole mass spectrometer and absolute quantitation is determined by comparing the area ratio of Heavy Internal Standard to Analyte signal. To quantitate low abundance proteins, strong cation exchange chromatography pre-fractionation can be performed on extremely complex peptide mixtures from samples such as tryptic digests of plasma.