Preparation of monoclonal antibody to human augmenter of liver regeneration:screening of hybridomas with unpurified antigen expressed by E.coli / 中国免疫学杂志

ABSTRACT

Objective:To prepare monoclonal antibody(McAb) against human augmenter of liver regeneration(hALR) by hybridoma technique with unpurified recombinant protein expressed by E.coli as screening antigen.Methods:BALB/c mice were immunized with hALR thioredoxin(hALR Thi) fusion protein twice.The splenocytes were fused with myeloma cells by polyethylene glycol and the hybridomas were selected in HAT medium.The positive hybridomas secreting anti hALR McAb were screened by means of ELISA,with the lysates of E.coli containing the recombinant plasmid pQE30 hALR induced by IPTG as screening antigen and containing plasmid pQE30 as controlling antigen.The reactivity of McAb secreted by the positive hybridoma to hALR expressed in eukaryocyte and native hALR in human serum was confirmed by means of ELISA and Western blot methods.Results:A positive hybridoma was screened successfully;the specific antigen antibody reaction was confirmed between the McAb prepared and hALR expressed in eukaryocyte and native hALR in human serum.Conclusion:Without any purification procedure,recombinant protein expressed by E.coli can be used for screening bybridomas in preparing of McAb with the expressed product of empty plasmid as control;anti hALR McAb can be used to the further study of hALR.