Poster 13.  Congresso da Sociedade Brasileira de Toxinologia, 8., Symposium of the Pan American Section of the International Society on Toxinology, 8., 2004, Angra dos Reis, Brasil.  Abstracts...  J. Venom. Anim. Toxins incl.Trop. Dis., 2004, 10, 3, p.372.

 

 

Rabbit Antisera Against Crotoxin and Phospholipase A2 (PLA2) From Crotalus durissus cascavella Neutralize the Enzymatic Activity of Crotoxin and PLA2 From This Venom

 

Beghini, D.G.1, Rodrigues-Simioni, L.2, Novello, J.C.1, Hyslop, S.2, Marangoni, S.1

 

1 Depto. de Bioquímica, IB, and 2 Depto. de Farmacologia, FCM, UNICAMP, Campinas, SP.

 

Crotoxin, the principal neurotoxin in the venom of Crotalusdurissus cascavella, contains a basic phospholipase A2 (PLA2) and an acidic protein, called crotapotin. The aim of this work was to examine the ability of antisera raised in rabbits against crotoxin (anti-Crtx) and PLA2 (anti-PLA2) in neutralizing the enzymatic activity of crotoxin and PLA2 from C. d. cascavella venom.

Rabbits were immunized with crotoxin and PLA2 purified from C. d. cascavella venom. The antibody titer during immunization was monitored by ELISA. Indirect hemolysis was used to assess the ability of the antisera to neutralize PLA2 activity. Initially, the HC50 (toxin concentration that produced ~50% of the maximum hemolysis) was determined. Neutralization was assayed by incubating HC50 of crotoxin or PLA2 with antiserum at a ratio of 1:1, for 30 min at 37°C.

The ELISA showed end-point dilutions of 3x10-6 for both antisera. The HC50 of the crotoxin and PLA2 was of 0.1 mg/ml and caused hemolytic haloes of  0.8±0.03 cm (n=8) and 0.85±0.02 cm (n=6), respectively. Anti-Crtx and anti-PLA2 fully neutralized the enzymatic HC50 activity of the crotoxin and PLA2. These results suggest interaction of the present antibodies in both antisera with “catalytic site” involved in the PLA2 activity.

 

Financial support: FAPESP

 

Correspondence to: beghini@unicamp.br