Purification Of Recombinant Schistosoma Japonicum Protein SjGST And Preparation And Characterization Of The Monoclonal Antibody

Objective To isolate and purify recombinant Schistosomiasis japonicum(Chinese strain) protein Glutathione S-transferase(GST)for preparing the monoclonal antibodies (McAbs) against rSjGST, We identified the biology characteristics of Schistosomiasis japonicum protein Glutathione S-transferase (rSjGST) and established the approach to detect the serum rSjGST levers of the patients. Methods The engineering Escherichia coli (E.coli) bacteria BL21 which contain the recombinant plasmid pET28a-rSjGST were cultured in the Lysogeny broth (LB) medium and induced with Isopropy-β-D-thiogalactoside (IPTG). The bacteria lysis contained the protein of interest (POI) with His-tag was then purified by affinity chromatograph and high performance liquid chromatography (HPLC). The immunological activity of recombined rSjGST after the two-step purification was identified in our research with SDS-PAGE. The female Balb/c mice were immunized with purified rSjGST, then the polyethylene glycol (PEG)-mediated fusion was conducted, the MCAbs were bolting by limiting dilution with indirect enzyme-linked immunosorbant assay (ELISA), and the hybridoma cell's chromosome were analysed by colchicine;the subtype of the MCAbs were identified with ISO-2 Mouse Monoclonal Antibody Isotyping Reagents, and the potencies of each McAb was detected.We established there approachs to detect the serum SjGST levels of the patients with Sandwich-ELISA, indirect-ELISA, Sandwich-ELISA combine with indirect-ELISA, estimated the specificity, sensitivity and Youden index of the approach. Results The data has shown that high purified rSjGST protein and the hybridoma cell lines 1B11 and 3C12 secreting monoclonal antibodies(McAbs) against rSjGST protein were obtained.The subtype of each McAb was confirmed to be IgG2b or IgG1. Conclusion The rSjGST protein is induced to express by isopropy-β-D-thiogalactoside (IPTG) in prokaryotic hosts Escherichia coli (E.coli) with high efficiency.The hybridoma cell lines 1B11 and 3C12 secreting monoclonal antibodies (McAbs) against rSjGST protein were harvested. They can be applied to further study on immunodiagnostic methods of Schistosoma japonicum.The approach to detect the serum SjGST levels of Schistosoma japonicum patients with Sandwich-ELISA combine with indirect-ELISA could be applied to clinical application as well.