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Development of an Antigen Spot Test for Detection of Coronavirus in Bovine Fecal Samples

机译：用于检测牛粪便样品中冠状病毒的抗原斑点试验的开发

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We have developed a rapid and sensitive microimmunodot blot assay, the antigen spot test (AST), for the detection of bovine coronavirus (BCV) antigen from neonatal calf fecal samples. The AST procedure can be completed in 3.5 h, whereas the previously reported immunodot blot assays require 10 to 12 h. Ninety-six samples can be tested per membrane, and 10 membranes (960 samples) may be processed by a single technologist in 1 working day. The effects of detergents, oxidizing chemicals, chaotropic agents, and enzyme substrates in improving the sensitivity and signal-to-noise ratio of the AST were studied. Finally, the sensitivity and specificity of AST for the detection of BCV antigen were compared to those of a sandwich enzyme-linked immunosorbant assay (ELISA) and a hemagglutination assay (HA). Of 347 field samples tested by all three methods, 94.2% were positive by AST, 91.4% were positive by ELISA, and 86.7% were positive by HA. The sensitivity of the AST was determined to be 100% compared to the results of the ELISA reference method. The specificity of the AST was 67%, which reflects a lower limit of detection of 104 viral particles per ml in a 10% fecal suspension.
机译：我们已经开发了一种快速灵敏的微量免疫印迹试验，即抗原斑点试验（AST），用于从新生儿小牛粪便样品中检测牛冠状病毒（BCV）抗原。 AST程序可以在3.5小时内完成，而先前报道的免疫斑点印迹分析需要10到12小时。每个膜可以测试96个样品，一个技术人员可以在1个工作日内处理10个膜（960个样品）。研究了去污剂，氧化剂，离液剂和酶底物在改善AST的灵敏度和信噪比方面的作用。最后，将AST检测BCV抗原的敏感性和特异性与夹心酶联免疫吸附测定（ELISA）和血凝测定（HA）的敏感性和特异性进行了比较。在这三种方法测试的347个野外样品中，AST呈阳性的占94.2％，ELISA呈阳性的占91.4％，HA呈阳性的占86.7％。与ELISA参考方法的结果相比，AST的灵敏度确定为100％。 AST的特异性为67％，这反映了在10％的粪便悬浮液中每毫升检测10 4 病毒颗粒的下限。

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