摘要
为解决目前蛇毒成份检测困难以及寻找一种可用于蛇伤快速鉴别诊断的方法,本研究建立了酶标抗原火箭免疫电泳法。本法结合了酶标法灵敏度高和火箭电泳操作简单的特点,对待测的同种抗原标记辣根过氧化物酶(酶标抗原),检测时把微量的酶标记抗原掺入被测样品中,在含多价抗血清的免疫电泳板进行电泳。最后用辣根过氧化物底物3.3′-二氨基联苯胺在板上直接显色,显色后观察到的免疫沉淀线高度与被测抗原含量呈正相关(r=+0.98)。采用本方法检测眼镜蛇毒细胞毒素和限镜王蛇毒 L-氨基酶氧化酶,细胞毒素的最低检测浓度为110ng/ml,氨基酸氧化酶为60ng/ml。比单向火箭免疫电泳法灵敏度高30倍。用同一原理的酶标记抗原融合电泳法监测L-氨基酸氧化酶的层析分离时,比酶活性法检测灵敏度高20倍。全部检测只需30分钟。初步观察商品眼镜蛇毒抗血清可对蛇毒15种成分产生免疫沉淀线,眼镜蛇的细胞毒与同科异科蛇毒的细胞毒素无交叉免疫反应。因此,采用本法可以确定不同蛇种蛇毒中所含的特异性抗原,获得蛇伤鉴别诊断的依据。
A noval immunoassay named Antigen-Enzyme Labelled Immnuno-electrophoresis (AELIE)with charccteristies of high specificity and sensityvity was developed.A homologous antigen was lablled with horseradish peroxidase,trace amout of this labessed antigen was mixed with sample and electrophoresis was perfomcd on an immunoplate containing ployspecifie antiserum.Rocket-shape immunoprccipitates containing ployspecifie antiserum.Rocket-shape immuno-preipitates were visualizde by incubation with enzyme substrate.Positive correlation(r=+0.98)was obtained between rocket hights and concentrations of antigen.In this study the method was applied for quantitation of eardiotoxin and L-amino acid oxidase in snake venoms,the sensitivity was 30 fold increase as compared with protein stain technique and 20 fold in- crease as compared with enzyme assay while monitoring chromatography.By distinqushing specific antigens in snake venoms this method may provide authentic evidence for diagnosis of snakebite and replace bicassay of various compounents in snake venoms.
出处
《蛇志》
1993年第2期10-14,共5页
Journal of Snake
关键词
免疫电泳
蛇毒
抗原
测定
Immunoelectrophoresis
Snake venom
Cardiotoxin
L-amino acid oxidase
