High speed digital protein interaction analysis using microfluidic single molecule detection system.

Abstract

The understanding of protein interaction dynamics is important for signal transduction research but current available techniques prove difficult in addressing this issue. Thus, using the microfluidic approach, we developed a digital protein analytical platform and methodology named MAPS (Microfluidic system Analyzing Protein in Single complex) that can measure the amount of target proteins and protein complexes at the digitally single molecule resolution. By counting protein events individually, this system can provide rough protein interaction ratios which will be critical for understanding signal transduction dynamics. In addition, this system only requires less than an hour to characterize the target protein sample, which is much quicker than conventional approaches. As a proof of concept, we have determined the interaction ratios of oncogenic signaling protein complexes EGFR/Src and EGFR/STAT3 before and after EGF ligand stimulation. To the best of our knowledge, this is the first time that the interaction ratio between EGFR and its downstream proteins has been characterized. The information from MAPS will be critical for the study of protein signal transduction quantitation and dynamics.