Aim: To construct and express the scFv phage antibody from natural anti-keratin monoclonal antibody 3B4 in E.coli and then compare its antigenic specificity with that of the parental antibody 3B4.
Methods: The V(H) and V(L) of 3B4 were amplified by PCR from pMD-18T- mAb 3B4 V(H) and pMD-18T- mAb 3B4 V(L), and then cloned into the phagemid pscMH. E.coli was transfected by recombined pscMH and superinfected by the helper phage VCSM13. Then the antigenic specificity of the scFv concentrated from the supernatant of the transfected E.coli was detected by indirect ELISA.
Results: DNA sequence analysis showed that the scFv sequence was the same as the original sequence except that there was one base mutation in the FR1 of V(L). Indirect ELISA showed that the scFv phage antibody was polyreactive as mAb 3B4, but its antigenic specificity was not the same with that of mAb 3B4.
Conclusion: The scFv phage antibody from natural anti-keratin monoclonal antibody 3B4 was successfully constructed and expressed. Its antigenic specificity was different from that of its parental antibody 3B4.