Application of a temperature-controllable microreactor to simple and rapid protein identification using MALDI-TOF MS

Abstract

We have carried out a simultaneous thermal denaturation and trypsin digestion of proteins using a temperature-controllable microreactor. This is a simple and rapid sample preparation technique for use before matrix-assisted laser desorption ionization time-of-flight mass spectrometry. In contrast to a conventional sample preparation method, which involves several chemical treatments, our sample preparation was performed using only trypsin digestion with the thermal denaturation of the target protein. Optimization of the reactor operational parameters for trypsin digestion using a temperature-controllable microreactor was carried out. The entire trypsin digestion procedure took about 11 min, and consisted of 1 min for the thermal denaturation of the sample protein (3 µl, 0.2 µM) at 85 °C, and 10 min for digestion of the protein at 37 °C. The resulting sequence coverage ranged from 24% to 57%, which was sufficient for practical protein identification.