Immunochemical study on beta1-bungarotoxin using polyclonal and monoclonal antibodies.

The number of antigenic determinants of beta1-bungarotoxin (beta1-Bgt), A chain and B chain were determined to be seven, five and two, respectively, by quantitative precipitin reactions and analysis of the soluble complex formed from beta1-Bgt and Fab fragments of the antibody. The gel filtration patterns on a size exclusion column revealed that the soluble complexes formed from non-precipitating antibody and beta1-Bgt at a different molar ratio all emerged in the void volume, indicating that the molecular weight of the soluble complex is around 6000 kDa or more. The gel filtration pattern from Fab fragments of non-precipitating antibody also revealed that only 1 or 2 molecules of Fab fragments are bound to beta1-Bgt, A chain and B chain instead of seven, five and two molecules as in the case of precipitating antibody. Besides, twenty-three stable monoclonal antibodies (mAbs) were prepared against beta1-Bgt by the hybridoma technique. Most of these mAbs cross-reacted with isotoxins of the beta-Bgt family, as determined by an enzyme-linked immunosorbent assay, but none of them reacted with notexin, notechis II-5 and three PLA2 homologues, PLA2, DE-I and CMS-9.