Sullender W M, Anderson L J, Anderson K, Wertz G W
Department of Microbiology, University of Alabama School of Medicine, Birmingham 35294.
J Clin Microbiol. 1990 Aug;28(8):1683-7. doi: 10.1128/jcm.28.8.1683-1687.1990.
A new approach to respiratory syncytial (RS) virus subgroup determination was developed by using a simple nucleic acid filter hybridization technique. By this method, virus-infected cells are bound and fixed in a single step, and the viral RNA in the fixed-cell preparation is characterized directly by its ability to hybridize to cDNA probes specific for either the A or B subgroups of RS virus. The subgroup-specific probes were constructed from cDNA clones that corresponded to a portion of the extracellular domain of the RS virus G protein of either a subgroup B RS virus (8/60) or a subgroup A RS virus (A2). The cDNA probes were labeled with 32P and used to analyze RS virus isolates collected over a period of three decades. Replicate templates of infected cell preparations were hybridized with either the subgroup A or B probe. The subgroup assignments of 40 viruses tested by nucleic acid hybridization were in agreement with the results of subgroup determinations based on their reactivities with monoclonal antibodies, which previously has been the only method available for determining the subgroup classification of RS virus isolates. The nucleic acid hybridization assay has the advantage of providing broad-based discrimination of the two subgroups on the basis of nucleic acid homology, irrespective of minor antigenic differences that are detected in assays in which monoclonal antibodies are used. The nucleic acid hybridization technique provides a reliable method for RS virus subgroup characterization.
通过使用一种简单的核酸滤膜杂交技术，开发出了一种呼吸道合胞（RS）病毒亚组测定的新方法。通过这种方法，病毒感染的细胞在一步操作中被结合并固定，固定细胞制剂中的病毒RNA通过其与RS病毒A或B亚组特异性cDNA探针杂交的能力直接进行表征。亚组特异性探针由对应于B亚组RS病毒（8/60）或A亚组RS病毒（A2）的RS病毒G蛋白细胞外结构域一部分的cDNA克隆构建而成。cDNA探针用32P标记，并用于分析在三十年时间内收集的RS病毒分离株。感染细胞制剂的重复模板与A或B亚组探针杂交。通过核酸杂交检测的40种病毒的亚组归属与基于它们与单克隆抗体反应性的亚组测定结果一致，而单克隆抗体反应性此前一直是确定RS病毒分离株亚组分类的唯一可用方法。核酸杂交测定法的优点是基于核酸同源性对两个亚组进行广泛区分，而不考虑在使用单克隆抗体的测定中检测到的微小抗原差异。核酸杂交技术为RS病毒亚组表征提供了一种可靠的方法。
