Generation of antiidiotypic antibody to monoclonal antibody MGd1 directed against gastric carcinoma by phage antibody library technique / 第三军医大学学报

ABSTRACT

Objective To pave the way for developing recombinant antiidiotypic antibody(antiId)vaccine of gastric carcinoma by generating phagedisplayed antiId to monoclonal antibody MGd1 directed against the cancer. Methods Balb/c mice were immunized i.p. with MGd1 conjugated with KLH, and mRNA was isolated from the spleens of the immunized mice. VH and VL DNAs of the antibody were amplified separately by RTPCR and assembled into ScFv DNAs with a linker DNA by PCR. The ScFv DNAs were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into competent E.coli TG1. The transformed cells were infected with M13KO7 helper phage to yield phage antibody ScFv library. After four rounds of panning to the library with MGd1, the MGd1positive clones were selected by ELISA from the enriched phages. The types of the antiId ScFv displayed on the selected phage clones were preliminary identified by competition ELISA. Results The VH,VL and ScFv DNAs were about 340 bp, 320 bp and 750 bp respectively. After four rounds of panning to the antibody library, 17 MGd1positive phage clones displayed antiId ScFv were selected from 40 enriched phage clones, among which 3 displayed ? or ? type antiId ScFv. Conclusion The successful generation of antiId ScFv to monoclonal antibody MGd1 by recombinant phage antibody library technology might lay a foundation for screening of novel candidate molecules for developing recombinant antiId vaccine of gastric carcinoma.