Anderson John P, Rascoe Lisa N, Levert Keith, Chastain Holly M, Reed Matthew S, Rivera Hilda N, McAuliffe Isabel, Zhan Bin, Wiegand Ryan E, Hotez Peter J, Wilkins Patricia P, Pohl Jan, Handali Sukwan
Division of Parasitic Diseases and Malaria, Centers for Disease Control and Prevention, Atlanta, Georgia, United States of America.
National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, United States of America.
PLoS Negl Trop Dis. 2015 Oct 20;9(10):e0004168. doi: 10.1371/journal.pntd.0004168. eCollection 2015.
The clinical spectrum of human disease caused by the roundworms Toxocara canis and Toxocara cati ranges from visceral and ocular larva migrans to covert toxocariasis. The parasite is not typically recovered in affected tissues, so detection of parasite-specific antibodies is usually necessary for establishing a diagnosis. The most reliable immunodiagnostic methods use the Toxocara excretory-secretory antigens (TES-Ag) in ELISA formats to detect Toxocara-specific antibodies. To eliminate the need for native parasite materials, we identified and purified immunodiagnostic antigens using 2D gel electrophoresis followed by electrospray ionization mass spectrometry. Three predominant immunoreactive proteins were found in the TES; all three had been previously described in the literature: Tc-CTL-1, Tc-TES-26, and Tc-MUC-3. We generated Escherichia coli expressed recombinant proteins for evaluation in Luminex based immunoassays. We were unable to produce a functional assay with the Tc-MUC-3 recombinant protein. Tc-CTL-1 and Tc-TES-26 were successfully coupled and tested using defined serum batteries. The use of both proteins together generated better results than if the proteins were used individually. The sensitivity and specificity of the assay for detecting visceral larval migrans using Tc-CTL-1 plus Tc-TES-26 was 99% and 94%, respectively; the sensitivity for detecting ocular larval migrans was 64%. The combined performance of the new assay was superior to the currently available EIA and could potentially be employed to replace current assays that rely on native TES-Ag.
由犬弓首蛔虫和猫弓首蛔虫引起的人类疾病临床谱，范围从内脏幼虫移行症和眼幼虫移行症到隐匿型弓首蛔虫病。寄生虫通常不会在受影响的组织中被检出，因此检测寄生虫特异性抗体通常是确诊所必需的。最可靠的免疫诊断方法是在酶联免疫吸附测定（ELISA）形式中使用弓首蛔虫排泄分泌抗原（TES-Ag）来检测弓首蛔虫特异性抗体。为了消除对天然寄生虫材料的需求，我们使用二维凝胶电泳，随后进行电喷雾电离质谱，鉴定并纯化了免疫诊断抗原。在TES中发现了三种主要的免疫反应性蛋白；所有这三种蛋白此前在文献中均有描述：Tc-CTL-1、Tc-TES-26和Tc-MUC-3。我们制备了大肠杆菌表达的重组蛋白，用于基于Luminex的免疫测定评估。我们无法用Tc-MUC-3重组蛋白建立功能性测定。Tc-CTL-1和Tc-TES-26成功偶联，并使用特定血清组进行了测试。两种蛋白一起使用产生的结果比单独使用时更好。使用Tc-CTL-1加Tc-TES-26检测内脏幼虫移行症的测定的灵敏度和特异性分别为99%和94%；检测眼幼虫移行症的灵敏度为64%。新测定的综合性能优于目前可用的酶免疫测定（EIA），并且有可能用于替代目前依赖天然TES-Ag的测定。
