Division of Veterinary Infection Biology and Immunology, Research Center Borstel, Parkallee 22, 23845 Borstel, Germany.
Parasitol Res. 2010 Oct;107(5):1241-8. doi: 10.1007/s00436-010-1994-8. Epub 2010 Aug 3.
Several DNA-based and serological tests have been established for the detection of Theileria annulata infection, including polymerase chain reaction, reverse line blot and loop-mediated isothermal amplification, indirect enzyme-linked immunosorbent assay (ELISA), and competitive ELISA. In this study, we have applied knowledge from the development and application of a recombinant protein-based indirect ELISA and competitive ELISA to establish a rapid test for point-of-care diagnosis of T. annulata infection in the field to be used by the veterinarian. For the development of a lateral flow test, the recombinantly expressed T. annulata surface protein (TaSP) was applied as the test antigen and anti-TaSP antiserum as the control line. TaSP antigen conjugated to colloidal gold particles was used as the detection system for visualization at the test line for the binding of anti-TaSP antibody present in the serum of infected animals. The developed test specifically detected antibodies in the serum of animals experimentally infected with T. annulata and showed no cross-reactivity with serum from animals infected with other tested bovine pathogens (Trypanosoma brucei, Anaplasma marginale, Babesia bigemina, Babesia bovis, and Theileria parva). Testing of field samples was compared to results obtained by other serological tests, resulting in a sensitivity and specificity of 96.3% and 87.5% compared to indirect fluorescence antibody test, 98.7% and 81.8% compared to indirect ELISA, and 100% and 47.6% compared to competitive ELISA. In conclusion, a rapid test for the detection of T. annulata infection (T. annulata lateral flow device, Ta-LFD) has been developed, which is easy to perform, delivers results to be read by the naked eye within 10 min, and is suitable for the detection of infection in field samples.
已经建立了几种基于 DNA 和血清学的检测方法来检测环形泰勒虫感染，包括聚合酶链反应、反向线印迹和环介导等温扩增、间接酶联免疫吸附试验（ELISA）和竞争 ELISA。在本研究中，我们应用了基于重组蛋白的间接 ELISA 和竞争 ELISA 的开发和应用知识，为兽医在现场即时诊断环形泰勒虫感染建立了一种快速检测方法。为了开发横向流动检测，应用重组表达的环形泰勒虫表面蛋白（TaSP）作为检测抗原，抗 TaSP 抗血清作为控制线。TaSP 抗原与胶体金颗粒偶联，作为检测系统，用于在测试线上可视化，用于检测感染动物血清中存在的抗 TaSP 抗体的结合。开发的检测方法特异性地检测到实验感染环形泰勒虫动物的血清中的抗体，与感染其他测试牛病原体（布氏锥虫、边缘无浆体、双芽巴贝斯虫、牛巴贝斯虫和小泰勒虫）的动物的血清无交叉反应。与其他血清学检测方法相比，对现场样本的检测结果显示，与间接荧光抗体检测相比，敏感性和特异性分别为 96.3%和 87.5%，与间接 ELISA 相比，敏感性和特异性分别为 98.7%和 81.8%，与竞争 ELISA 相比，敏感性和特异性分别为 100%和 47.6%。总之，已经开发出一种用于检测环形泰勒虫感染的快速检测方法（环形泰勒虫横向流动装置，Ta-LFD），该方法易于操作，在 10 分钟内即可通过肉眼读取结果，适用于现场样本的检测。
