3B-O2-1625 PDF
抗体のバイオアナリシスに向けた安定同位体標識trastuzumab Fabの発現とLC/MS/MSによる高感度定量
The LC/MS/MS based bioanalysis of biologics attracts attention as a technology complementing the conventional Ligand Binding Assay. In general, because of the cost efficiency, synthetic isotope labeled peptides derived from the target protein have been used as internal standard in the LC/MS/MS based protein quantification instead of the protein itself. In this paper, we will report the preparation and the utilization of the stable isotope labeled trastuzumab Fab with using of the Corynex® (C. glutamicum protein expression system) technology, which is relatively affordable, for the LC/MS/MS based bioanalysis. In consequence, the prepared labeled trastuzumab was 15N >98% isotope purity by intact MS and peptide mapping analysis, and no differences between the synthetically prepared isotope labeled peptide in the parallel reaction monitoring (PRM) quantitative assay. Also, stable isotope labeled trastuzumab Fab enabled to add the target peptides easily without any further preparations.