REACTIVITY OF MONOCLONAL ANTIBODIES ANTI-K49 MYOTOXINS WITH VENOMS FROM DIFFERENT SNAKE FAMILIES

BENTO, L., TANJONI, I., SPENCER, P.J., FERNANDES, I., MOURA-DA-SILVA, A.M.

Laboratório de Imunopatologia, Instituto Butantan, São Paulo, Brasil

Myotoxins are components widespread in venoms of snakes from different genera, responsible for muscle tissue damage adjacent to the bite.  These toxins belong to two major structural groups: short chain peptides and molecules with a phospholipase A2 structure with preserved catalytic activity (D-49) or a mutated non-catalytic form (K-49). Monoclonal antibodies (MoAb) were previously raised against Bothrops jararacussu K-49 Bothropstoxin-1. In this work, we analyzed their reactivity against isolated toxin, homologous venom and venoms of 27 different species of snakes. MoAbs producing cells were expanded in vitro and subsequently injected into mice to produce the ascites. Antibody levels were assayed by ELISA. Similar titres were detected against isolated toxin or whole venom, although differing amongst the clones. When analyzed by Western blotting, MoAbs recognized a B. jararacussu venom band of approximately 28 kDa under non-reducing conditions and a 14 kDa band in reduced samples corresponding to the dimeric and monomeric forms of the myotoxin, respectively. Four antibodies reacted with reduced samples, three reacted preferentially with non-reduced samples and two were non-reactive. MoAbs were then tested by dot blot, under native conditions with venoms of 27 species of snakes belonging to VIPERIDAE, ELAPIDAE and COLUBRIDAE families. MoAbs reacted preferentially with venoms of viper snakes, sub-family CROTALINAE, preferentially with Bothrops jararacussu, Bothrops jararaca, Bothrops asper,

Crotalus atrox, Crotalus adamanteus and Agkistrodon contortrix. Neutralization ability of MoAbs and their reactivity towards K-49 or D-49 myotoxins are under evaluation. Concluding, these antibodies are apparently selective to myotoxins, being an interesting tool for screening of venoms and purification of toxins by immunoaffinity.

Financial support: FAPESP and CNPq (PIBIC fellowship for L.B.)

CORRESPONDENCE TO:

MOURA-DA-SILVA, A.M., Laboratório de Imunopatologia, Instituto Butantan, 05503-900, São Paulo, Brasil, Email: anamoura@usp.br