Protection against dermonecrotic and lethal activities of Loxosceles intermedia spider venom by immunization with a fused recombinant protein
Protection against dermonecrotic and lethal activities of Loxosceles intermedia spider venom by immunization with a fused recombinant protein
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DOI：
10.1016/s0041-0101(02)00282-9
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发表时间：
2003-03-01
期刊：
影响因子：
2.8
通讯作者：
Chávez-Olórtegui, C
中科院分区：
文献类型：
--
作者：
Araujo, SC;Castanheira, P;Chávez-Olórtegui, C
我们报告了使用重组中间剑水蚤蜘蛛蛋白的融合蛋白的形式作为免疫兔和小鼠的抗原。我们的目的是在这些动物中产生模型保护性抗血清，以对抗来自巴西蜘蛛的毒液的皮肤坏死和致死活性，巴西蜘蛛每年报告3000例蜘蛛咬伤。从L. intermedia毒腺，在E.大肠杆菌细胞作为与β-半乳糖苷酶的融合蛋白，并通过分子过滤和亲和层析纯化重组蛋白（Li-rec蛋白）[Kalapothakis等，Toxicon（2002）in press].对Li-rec蛋白进行了表征，并将其用作抗原以在兔和小鼠中产生抗体。这些特异性产生的抗体识别天然毒液。体外中和实验表明，1 ml抗Li-rec蛋白的兔血清能中和25 LD_（50）的全毒。在体内保护实验中，融合蛋白诱导兔对天然毒液的皮肤坏死活性的长期保护。用不同剂量的Loxosceles毒液攻击免疫小鼠。小鼠完全免受2.5 LD 50毒液的伤害。这一结果提供了使用这种重组蜘蛛蛋白作为免疫原的抗蛇毒血清的临床应用或可用作疫苗提供有效的免疫保护中间型乳杆菌毒液的发展的基础数据。(C)2003爱思唯尔科技有限公司版权所有。
We report the use of a recombinant Loxosceles intermedia spider protein in the form of a fusion protein as an antigen for immunization in rabbits and mice. The aim is to produce model protective antisera in these animals against dermonecrotic and lethal activities of the venom from the Brazilian spider responsible for 3000 cases, reported annually, of spider bites in South Brazil. A protein homologous to the dermonecrotic toxin was cloned from a cDNA expression library made with L. intermedia venom glands, expressed in E. coli cells as a fusion protein with beta-galactosidase and the recombinant protein (Li-rec protein) was purified by molecular filtration and affinity chromatography [Kalapothakis et al., Toxicon (2002) in press]. The Li-rec protein was characterized and used as an antigen to generate antibodies in rabbits and mice. These specifically raised antibodies recognized the native venom. In vitro neutralization assay of lethal effects indicated that 1 ml of rabbit serum raised against Li-rec protein was able to neutralize 25 LD50 of the whole venom. In vivo protection experiments, the fusion proteins induced a long-term protection in rabbits against the dermonecrotic activity of the native venom. Immunized mice were challenged with various doses of the Loxosceles venom. Mice were fully protected against 2.5 LD50 of venom. This result provides basic data for the use of such recombinant spider proteins as immunogens in the development of anti-venoms for clinical use or can be used as a vaccine providing efficient immune protection against L intermedia venom. (C) 2003 Elsevier Science Ltd. All rights reserved.