Multiple reaction monitoring for absolute quantification

In multiple reaction monitoring (MRM) using triple quadrupole (/linear ion trap) instruments the quadrupole 1 (Q1) is set for a distinct precursor mass that enters the collision chamber. In contrast to product ion scanning where all the fragments are scanning through the third quadrupole (Q3), Q3 is set constant for one or more distinct fragment masses. In this manner the transition Q1 – Q3 is monitored. The signal is highly specific and is thus used to detect distinct protein in a very complex mixture. Moreover its intensity is proportional to sample amount over five orders of magnitude. Some approaches uses MRM in a hypothesis driven manner to identify post-translational modifications in a protein of interest (MIDAS).

We apply MRM for absolute protein quantification with the aim to elucidate the stoichiometry of single proteins in protein (-RNA) complexes by using labelled AQUA peptides (Link) that are chosen by proteomtypic peptide criteria.