Figure 4
Prokaryotic expression and purification of rERp29, and production of polyclonal anti-rERp29 antibodies in rabbits. (A) Expression and purification of rERp29 fragment in E. coli BL21 (DE3) host cells. Different fractions of E. coli were separated on a Coomassie-blue-stained 15% SDS-PAGE gel. Lane M, protein molecular weight markers; lane 1, total cell lysates carrying pET-28a(+)/ERp29 fragment before IPTG induction; lane 2, total cellular protein after IPTG induction; lane 3, supernatant of cell lysate after ultrasonic treatment; lane 4, pellet of cell lysate after ultrasonic treatment; lane 5, protein purified based on its His6-tag by affinity chromatography employing a Ni2+-NTA His-binding resin; lane 6, protein eluted in PBS by cutting certain protein bands from the gel after preparative electrophoresis. (B) Values of ELISA absorbance at 450 nm using rERp29 protein immunized rabbit sera or pre-immune serum (control).