Identification of rhPDGF-BB isoforms.

Abstract

<p>(A) Five isoforms detected by SDS-PAGE were transferred to a PVDF membrane for protein N-terminal sequencing. (B) Schematic representation of N-terminal sequencing results. Bands I, II, and III represent intact PDGF-BB, while bands IV and V are truncated isoforms generated by the cleavage at the Arg 27-Thr 28 site. (C) rhPDGF-BB (5 μg) was subjected to SDS-PAGE under reducing conditions and analyzed by western blotting, Coomassie Blue staining, and glycosylation staining, respectively. Two bands corresponding to isoforms III and IV stained by Coomassie Blue could be detected by antibody staining (marked with red lines). Glycosylation staining revealed another two bands corresponding to isoforms I and II (marked with red lines). (D) SDS-PAGE and subsequent glycosylation staining of higher rhPDGF-BB load (10 μg and 20 μg) revealed three bands; the third band with the lowest molecular weight was assumed to correspond to isoform IV.</p