Departamento de Medicina Molecular y Bioprocesos, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Av. Universidad 2001, Colonia Chamilpa, Cuernavaca, Morelos, C.P. 62210, Mexico.
National Natural Toxins Research Center (NNTRC) and Department of Chemistry, Texas A&M University-Kingsville, Kingsville, TX, USA.
Toxicon. 2021 Jul 15;197:70-78. doi: 10.1016/j.toxicon.2021.04.005. Epub 2021 Apr 21.
Crotamine is a paralyzing toxin (MW: ~5 kDa) found in different proportions in some rattlesnake venoms (up to 62%). Mexican pit viper antivenoms have shown low immunoreactivity against crotamine, which is an urgent quality to be improved. The objective of this work was to evaluate the ability of a novel recombinant fusion protein composed of sphingomyelinase D and crotamine, and two whole venoms from Crotalus molossus nigrescens and C. oreganus helleri to produce neutralizing antibodies against crotamine. These immunogens were separately used for immunization procedures in rabbits. Then, we generated three experimental antivenoms to test their cross-reactivity via western-blot against crotamine from 7 species (C. m. nigrescens, C. o. helleri, C. durissus terrificus, C. scutulatus salvini, C. basiliscus, C. culminatus and C. tzabcan). We also performed pre-incubation neutralization experiments in mice to measure the neutralizing potency of each antivenom against crotamine induced hind limb paralysis. Our antivenoms showed broad recognition across crotamine from most of the tested species. Also, neutralization against crotamine paralysis symptom was successfully achieved by our three antivenoms, albeit with different efficiencies. Our results highlight the use of crotamine enriched venoms and our novel recombinant fusion protein as promising immunogens to improve the neutralizing potency against crotamine for the improvement of Mexican antivenoms.
响尾蛇毒素是一种麻痹毒素（MW：~5 kDa），在某些响尾蛇毒液中以不同比例存在（高达 62%）。墨西哥响尾蛇抗蛇毒血清对响尾蛇毒素的免疫反应性较低，这是一个亟待改进的质量问题。本工作的目的是评估由鞘磷脂酶 D 和响尾蛇毒素组成的新型重组融合蛋白，以及来自 Crotalus molossus nigrescens 和 C. oreganus helleri 的两种全毒液产生针对响尾蛇毒素的中和抗体的能力。这些免疫原分别用于兔的免疫程序。然后，我们生成了三种实验性抗蛇毒血清，以通过 Western-blot 测试它们对来自 7 种（C. m. nigrescens、C. o. helleri、C. durissus terrificus、C. scutulatus salvini、C. basiliscus、C. culminatus 和 C. tzabcan）响尾蛇毒素的交叉反应性。我们还在小鼠中进行了预孵育中和实验，以测量每种抗蛇毒血清对响尾蛇毒素诱导的后肢瘫痪的中和效力。我们的抗蛇毒血清显示出对大多数测试物种的响尾蛇毒素的广泛识别。此外，我们的三种抗蛇毒血清成功地实现了对响尾蛇毒素麻痹症状的中和，尽管效率不同。我们的结果强调了使用富含响尾蛇毒素的毒液和我们的新型重组融合蛋白作为有前途的免疫原，以提高对响尾蛇毒素的中和效力，从而改善墨西哥抗蛇毒血清。
