Beghini Daniela G, Hernandez-Oliveira Saraguaci, Rodrigues-Simioni Léa, Novello José Camilo, Hyslop Stephen, Marangoni Sérgio
Departamento de Bioquímica, Instituto de Biologia, Universidade Estadual de Campinas (UNICAMP), CP 6109, CEP 13083-970 Campinas, SP, Brazil.
Toxicon. 2004 Aug;44(2):141-8. doi: 10.1016/j.toxicon.2004.05.011.
Crotoxin, the principal neurotoxin in venom of the South American rattlesnakes Crotalus durissus terrificus and Crotalus durissus cascavella, contains a basic phospholipase A2 (PLA2) and an acidic protein, crotapotin. In this work, we examined the ability of rabbit anti-sera against crotoxin and its PLA2 subunit to neutralize the neurotoxicity of venom and crotoxin from C. d. cascavella in mouse phrenic nerve-diaphragm and chick biventer cervicis preparations. Immunoblotting showed that the anti-sera recognized C. d. cascavella crotoxin and PLA2. This was confirmed by ELISA, with both anti-sera having end-point dilutions of 3 x 10(-6). Anti-crotoxin serum neutralized the neuromuscular blockade in phrenic nerve-diaphragm muscle preparations at venom or crotoxin:anti-serum ratios of 1:2 and 1:3, respectively. Anti-PLA2 serum also neutralized this neuromuscular activity at a venom or crotoxin:anti-serum ratio of 1:1. In biventer cervicis preparations, the corresponding ratio for anti-crotoxin serum was 1:3 for venom and crotoxin, and 1:1 and 1:2 for anti-PLA2 serum. The neutralizing capacity of the sera in mouse preparations was comparable to that of commercial anti-serum raised against C. d. terrificus venom. These results show that anti-sera against crotoxin and PLA2 from C. d. cascavella venom neutralized the neuromuscular blockade induced by venom and crotoxin in both nerve-muscle preparations, with the anti-serum against crotoxin being slightly less potent than that against crotoxin.
响尾蛇毒素是南美响尾蛇（Crotalus durissus terrificus和Crotalus durissus cascavella）毒液中的主要神经毒素，它包含一种碱性磷脂酶A2（PLA2）和一种酸性蛋白，即响尾蛇蛋白。在本研究中，我们检测了兔抗响尾蛇毒素血清及其PLA2亚基中和C. d. cascavella毒液和响尾蛇毒素对小鼠膈神经-膈肌及鸡颈二腹肌标本神经毒性的能力。免疫印迹显示抗血清能识别C. d. cascavella响尾蛇毒素和PLA2。酶联免疫吸附测定（ELISA）证实了这一点，两种抗血清的终点稀释度均为3×10⁻⁶。抗响尾蛇毒素血清在毒液或响尾蛇毒素与抗血清的比例分别为1:2和1:3时，能中和膈神经-膈肌肌肉标本中的神经肌肉阻滞。抗PLA2血清在毒液或响尾蛇毒素与抗血清的比例为1:1时也能中和这种神经肌肉活性。在颈二腹肌标本中，抗响尾蛇毒素血清对毒液和响尾蛇毒素的相应比例为1:3，抗PLA2血清的相应比例为1:1和1:2。小鼠标本中血清的中和能力与针对C. d. terrificus毒液产生的商业抗血清相当。这些结果表明，针对C. d. cascavella毒液中响尾蛇毒素和PLA2的抗血清能中和两种神经肌肉标本中由毒液和响尾蛇毒素诱导的神经肌肉阻滞，抗响尾蛇毒素血清的效力略低于抗PLA2血清。
