Fischer C, Nosratian R, Habtemichael N, Tolle R, Riemenschneider V, Geyer E
Fachbereich Biologie, Philipps-Universität Marburg, Germany.
Trop Med Parasitol. 1994 Dec;45(4):324-8.
A Taenia crassiceps metacestode cDNA expression library in lambda gt 11 was screened with rabbit antisera to metacestodal T. solium and T. saginata crude extract. Primary clones (121) were identified, and after rescreening and lysogenization in Escherichia coli Y 1089, were tested in Western blot for reactivity with the same antisera. In addition, analyses were performed with rabbit antisera directed towards T. crassiceps and Echinococcus granulosus metacestode crude extract, sera from humans with neurocysticercosis (Mexico) and other important helminth diseases, mice and calves with experimental T. crassiceps and T. saginata infections and normal sera. Of those tested, 22 clones expressing beta-galactosidase fusion proteins (approximately 118-132 kDa) were reactive with IgG antibodies of cysticercotic patients and T. crassiceps infected mice. Of these clones, 11 were also sero-positive with calf-IgG antibodies against T. saginata larvae. None of the 22 clones reacted with IgG antibodies due to human cystic and alveolar echinococcosis, intestinal/hepatic or urinary schistosomiasis, African onchocerciasis or with sera from uninfected controls (man, rabbit, calf and mouse). Of these 22 clones, 15 have been subcloned into the plasmid vectors pGEX-2T (modified) and pT7T3 alpha 19. Expressed glutathione-S-transferase fusion proteins were again tested for sensitivity and specificity by Western blot, and concentrated by affinity chromatography. The nucleotide sequence of the cDNA inserts of 9 clones has been determined in pT7T3 alpha 19 and revealed identity in 4 and 5 clones, respectively.
用兔抗猪带绦虫和牛带绦虫囊尾蚴粗提物的抗血清筛选λgt 11载体中的肥胖带绦虫囊尾蚴cDNA表达文库。鉴定出121个初级克隆，在大肠杆菌Y 1089中再次筛选并溶源化后，用Western印迹法检测其与相同抗血清的反应性。此外，还使用了针对肥胖带绦虫和细粒棘球绦虫囊尾蚴粗提物的兔抗血清、来自患有神经囊尾蚴病（墨西哥）和其他重要蠕虫病的人类的血清、感染实验性肥胖带绦虫和牛带绦虫的小鼠和小牛的血清以及正常血清进行分析。在检测的克隆中，22个表达β-半乳糖苷酶融合蛋白（约118 - 132 kDa）的克隆与囊尾蚴病患者和感染肥胖带绦虫的小鼠的IgG抗体发生反应。在这些克隆中，11个对牛抗牛带绦虫幼虫的IgG抗体也呈血清阳性。这22个克隆均未与因人类囊性和肺泡型棘球蚴病、肠道/肝脏或泌尿生殖道血吸虫病、非洲盘尾丝虫病产生的IgG抗体反应，也未与未感染对照（人、兔、小牛和小鼠）的血清反应。在这22个克隆中，15个已亚克隆到质粒载体pGEX - 2T（修饰型）和pT7T3α19中。通过Western印迹法再次检测表达的谷胱甘肽 - S - 转移酶融合蛋白的敏感性和特异性，并通过亲和层析进行浓缩。已确定pT7T3α19中9个克隆的cDNA插入片段的核苷酸序列，分别在4个和5个克隆中显示出一致性。
