Production and Characterization of Anti—estrone Monoclonal Antibody

Determination of estrone(E1)levels has a significant meaning in evaluating physiological effect and diagnsing some diseases,In order to detect free E1 in bilogical fluids,a monoclonal antibody specific for E1 was prepared after the complete antigen of E1 was synthesized,The purified monoclonal antibody was fully characterized for later immunoassay.Methods 3-O-carboxymethyl ether derivative of E1 was synthesized and in turn coupled to bovine serum albumin (BSA)to form complete antigen E1－BSA,A monocloanl antibody(McAb)specific for E1 was produced both in vitro and in vivo by a hybridoma anti-E1 Anti-E1was prepared by fusion of SP2/0 murine myeloma cells with spleen cells isolated from immunized BALB/c mouse.The McAb was characterized by enzyme-linked immunosorbent assay(ELISA),SDS-PAGE and Western-blotting THe specificity of the immunoassay was investigated by determining the cross-reastions of E1 analogs when free E1 was detected by competitive indirect enzyme-linke immounosorbent assay(CI-ELISA).Results Analysis revealed that anti-E1 McAb(E1-McAb)was of the IgG1 tppe,the molecular weight of E1-McAb was 164000 daltons,The affinity constant of E1-McAb with coated complete antigen was 8.2×10^8L/mol,The linear range for free E1 determined by CI-ELISA was 10pg/mL-10ng/mL,The detection limit was 21.4pg/mL(defined as twice the standard deviation of the blank),Conclusion The CI-ELISA developed with E1-McAb was both sersitive and specific.The prepared E1-McAb can be used in some immunoassays.