Protein expression analysis has become increasingly important in various biological research fields. The increased accuracy, reproducibility, and large dynamic range provided by selected reaction monitoring SRM technology suggest that SRM is likely to become the platform of choice for high-throughput and accurate expression profiling of selected proteins. One of the challenging, yet easily overlooked, aspects of the analysis of SRM data is how to normalise samples so that comparisons can be made across samples. Most approaches that utilise heavy isotope reference peptides to normalise endogenous samples work readily when the reference peptides are highly pure and have stable transitions; otherwise, the approaches become problematic. We demonstrate herein the normalisation problem that might frequently occur with impure reference peptides and propose a possible alternative strategy. The alternative strategy that we propose is the use of extra information, which can be obtained in the initial step of the SRM experiment. The simulation study showed that the proposed strategy successfully soften the normalisation problem arising from the impurity of standard, isotope-labelled peptides.
['Jungsoo Gim', 'Taesung Park']
The effect of impurity of reference peptides on normalisation in selected reaction monitoring experiments