Expression, purification and characterization of HSA-related recombinant proteins. (A) Electrophoresis of conditioned media from P. pastoris transformed with different expression plasmids. Conditioned media samples taken 24, 48, 72, or 96 hours after the start of methanol induction are shown on 10% reducing SDS-polyacrylamide gels stained with Coomassie Blue. The protein product encoded by the expression plasmid is shown above each gel. The position of molecular mass markers (200, 150, 120,. 100, 85, 70, 60, 50, 40, and 30 kDa) is shown to the left of each panel and labelled (XL4-HSA) in one instance as an example. (B) Upper panel, gel profile of purified recombinant proteins on 10% SDS-polyacrylamide gels electrophoresed under reducing conditions and stained with Coomassie Blue. Proteins (400 ng) are identified above each lane as in Figure 1A, plus GST (E.coli-expressed glutathione sulfotransferase); M, molecular mass markers identical to those used in Figure 1A. Middle and lower panel, replicate gels differing only in the amounts of protein loaded per lane, to avoid overloading with antibodies of different affinity, were immunoblotted and probed with Anti-HSA antibodies (middle pane, 100 ng/lane) or anti-hexahistidine antibodies (lower panel, Anti-His, 500 ng/lane).
Sheffield and Eltringham-Smith BMC Biotechnology 2011 11:127 doi:10.1186/1472-6750-11-127