SDS polyacrylamide gel electrophoresis showing (A) Coomassie brilliant blue stained 12% gel of proteins isolated from lane 1, noninduced culture of BL21DE3 plys S transformed with serpin 2A ORF cloned into the Pet 3A T7 RNA polymerase expression vector; lane 2, the same culture after 2 hours of induction at 37°C with 0.2 mmol/L IPTG and a Western immunoblot of a duplicate gel probed with the serpin 2A peptide antiserum. Detection was by ECL and the exposure time for the blot shown was 3 seconds. (B) Western immunoblot of proteins isolated from an FDCP-Mix G/M differentiation time course. The +ve control lane was made up of proteins isolated from the amphotropic retrovirus packaging cell line GP + envAm1213 that had been transfected with the serpin 2A ORF cloned into the high level retroviral expression vector M5. The −ve control lane was made up of proteins isolated from the wild type packaging cell line. This was incubated with the serpin 2A peptide antiserum with detection by peroxidase conjugated antiimmunoglobulin and ECL. Exposure times were typically between 1 and 2 minutes. (C) Western immunoblot of proteins isolated from Con A + Il-2 activation time course of primary splenocyte T cells. Lanes 0 to 5 represent the period of the activation in days. These were incubated with the serpin 2A antiserum and detection was by ECL with an exposure time of 1 minute.
