A DOUBLE SANDWICH MONOCLONAL ENZYME-IMMUNOASSAY FOR DETECTION OF HEPATITIS-B SURFACE-ANTIGEN

An enzyme immunoassay for detection of hepatitis B surface antigen bas ed on the use of 3 monoclonal antibodies (mAbs) was developed: an IgM as capture and 2 IgG1 for detection. The system biotin-streptavidin wa s compared with direct conjugation of mAbs to peroxidase and was prefe rred because of its higher signal to noise ratio. The possibility of s imultaneous addition of human serum and biotin-mAb was discarded becau se of an evident prozone effect with some sera containing high HBsAg l evels. The conjugation of biotin to IgG1 mAbs through a spacer arm (am idocaproil) and the use of a highly sensitive substrate (tetramethylbe nzydine) improved the assay detection limit by about 10 times.