间斑寇蛛L.tredecimguttatus粗毒抗血清制备及其活性分析

Preparation and Bioactivity Analysis of Antiserum Against Latrodectus tredecimguttantus Venom

间斑寇蛛Latrodectustredecimguttatus是世界上毒性最强的蜘蛛之一,其毒液中含有许多大分子物质,可用于制备抗血清。将粗毒过滤除菌后作为抗原,加弗氏佐剂背部皮下免疫豚鼠。加强免疫3次后经检测抗血清效价达到1:8,再次加强免疫后断颈处死豚鼠放血,分离血清,经饱和硫酸铵分级沉淀及阴离子交换柱层析纯化后,采用westernblot法检测抗体所在峰。离体大鼠输精管电刺激收缩反应实验及活体动物给药实验均表明抗体对毒素具有明显的中和作用。特异性间斑寇蛛毒素抗血清为治疗间斑寇蛛咬伤奠定了基础。

The spider Latrodectus tredecimguttantus is one of the most venomous spiders in the world. The venom of this spider contains many kinds of proteins which can be used to prepare antiserum. The filtered crude venom was mixed with Freund＇ s adjuvant and then injected subcutaneously into the back of guinea pig. After the third reimmunization, the titre of antiserum reached 1：8 determined through double immunodiffusion detection. The fourth reimmunization was performed and then the experimental animals were bled. Then the separated serum was purified by ammonium sulphate fractionation and anion-exchange chromatography. To confirm the antivenom, all of the eluted peaks were detected by western blot. The neutralization of the toxic effect ofL. tredecimguttatus venom by the antivenom was assessed in vitro with an isolated rat nerve-spermaduct preparation and in vivo with a lethality experiments in mice. Both experiments showed the obvious neutralization effects of the antivenom. The results demonstrated that we prepared a novel specific antivenom against L. tredecimguttatus venom successfully, and this made a foundation for the new specific treatment of the envenomation by the spider L. tredecimguttatus.