Construction of human-mouse hybrid Fab phage antibody libraries / 解放军医学杂志

ABSTRACT

Objective To construct human-mouse hybrid Fab phage antibody libraries and to select target Fab genes. Methods With Fd repertoire genes of PBMC from hepatoma patients and the chemeric light chain gene of cFab-HAb18, two hybrid Fab phage antibody libraries(pComb3 and pComb3X) were constructed. The GST fusion and non-fusion HAb18GE were used as antigens to select the target antibodies with the optimized strategies of amplifying and screening. Results When pComb3 displaying library was cultivated at 37℃, the target fragments could not be cut out with the restriction endonucleases from most clones of the library, which was an indication of “recombinant-deletion”. But no “recombinant-deletion” was found in pComb3X displaying library. Because of the serious cross-reaction, the phage-display Fab ELISA could not be used for the selection of positive clones. But positive clones could be identified in the supernatant of lysate with ELISA afterbeing induced by IPTG. Conclusion Through the construction of HuMFab phage antibody libraries and the optimization of screening strategies, we get the desired HuMFab genes for subsequent research.