Fig. 1

Overexpression and purification of recombinant hPLSCR1 in E. coli BL21 (DE3). a Schematic showing the steps involved in recombinant purification of hPLSCR1 b Coomassie stained SDS-PAGE gel showing hPLSCR1 at each stage of purification: Recombinant hPLSCR1 was overexpressed in E. coli BL21 (DE3) and seen as an intense band in lane 3, compared to the uninduced cells (Lane 2). The cells were then lysed by sonication. Most of the overexpressed protein was extracted as inclusion bodies (Lane 5) with low amounts in the soluble fraction (Lane 4). N-Lauroylsarcosine recovered active protein from the inclusion bodies into the soluble fraction (Lane 6). c Silver stained SDS-PAGE gel showing purified hPLSCR1: The N-LS recovered protein was purified to homogeneity by Ni²⁺ - NTA chromatography and the elutes were loaded on to 12 % SDS PAGE. d DEAE anion exchange chromatography was performed for the elutes from Ni²⁺-NTA chromatography and then eluted with a NaCl gradient. hPLSCR1 eluted in 350 mM NaCl (Lane 5–8). e The 350 mM DEAE fractions containing hPLSCR1 were pooled and again passed through Ni²⁺-NTA resin and eluted with 250 mM imidazole. The eluted samples were then concentrated and estimated. Left panel shows the silver stained SDS-PAGE gel showing 20 pmol and 200 pmol of purified hPLSCR1. Western blotting was performed for the purified hPLSCR1, where a specific monoclonal antibody against hPLSCR1 was used and visualized by chemiluminescence as shown in the right panel