Vitta Apichat, Yoshino Timothy P, Kalambaheti Thareerat, Komalamisra Chalit, Waikagul Jitra, Ruangsittichai Jiraporn, Dekumyoy Paron
Department of Helminthology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand.
Southeast Asian J Trop Med Public Health. 2010 Jul;41(4):785-99.
The aim of this study was to find novel proteins expressed from an Angiostrongylus cantonensis adult female worm cDNA library for serodiagnosis of angiostrongyliasis. An immuno-dominant clone, fAC22, was identified by immunoscreening with pooled positive sera from proven angiostrongyliasis patients. The clone contained an open reading frame of 2,136 bp encoding a 80.5 kDa protein with a predicted isoelectric point of 5.8. The deduced amino acid sequence (712 amino acids) contained the conserved domain of Small mutS related (Smr) superfamily protein, with similarity with the Smr domain protein of Brugia malayi. The fusion His-tagged 81 kDa recombinant protein expressed as inclusion body in Escherichia coli was solubilized and purified by Ni-affinity chromatography for use in immunoblot analysis. Its sensitivity, specificity, positive and negative predictive values in immunodiagnostic test was 93.5, 91.5, 79.0 and 97.5%, respectively. Although some cross-reactivity of the antigen was observed among gnathostomiasis, bancroftian filariasis, ascariasis, echinococcosis, paragonimiasis and opisthorchiasis, sera from 14 other infections were all negative. These data indicate its possible application in immunodiagnosis of clinically suspected angiostrongyliasis. Key words: Angiostrongylus cantonensis,eosinophilic meningitis, recombinant fusion protein, immunodiagnosis
本研究旨在从广州管圆线虫成虫雌性虫体cDNA文库中寻找新的蛋白质，用于广州管圆线虫病的血清学诊断。通过用确诊的广州管圆线虫病患者的混合阳性血清进行免疫筛选，鉴定出一个免疫显性克隆fAC22。该克隆包含一个2136 bp的开放阅读框，编码一个80.5 kDa的蛋白质，预测的等电点为5.8。推导的氨基酸序列（712个氨基酸）包含Small mutS相关（Smr）超家族蛋白的保守结构域，与马来布鲁线虫的Smr结构域蛋白相似。在大肠杆菌中以包涵体形式表达的融合His标签的81 kDa重组蛋白经Ni-亲和层析溶解和纯化，用于免疫印迹分析。其在免疫诊断试验中的敏感性、特异性、阳性和阴性预测值分别为93.5%、91.5%、79.0%和97.5%。尽管在颚口线虫病、班氏丝虫病、蛔虫病、棘球蚴病、肺吸虫病和华支睾吸虫病中观察到该抗原存在一些交叉反应，但来自其他14种感染的血清均为阴性。这些数据表明其在临床疑似广州管圆线虫病免疫诊断中的可能应用。关键词：广州管圆线虫；嗜酸性粒细胞性脑膜炎；重组融合蛋白；免疫诊断
