Cubie H A, Inglis J M, Leslie E E, Edmunds A T, Totapally B
Regional Virus Laboratory, City Hospital, Edinburgh, Scotland.
J Med Virol. 1992 Dec;38(4):283-7. doi: 10.1002/jmv.1890380410.
Direct detection of respiratory syncytial (RS) virus antigen in nasopharyngeal secretions (NPS) provides the most rapid diagnostic test for RS infection, but more sensitive methods might be more beneficial in the study of virus shedding. RS virus RNA was extracted from cells stored at -70 degrees C either in suspension with added RNAse inhibitor or as a pellet without inhibitor. The RNA was reverse transcribed, the resultant cDNA amplified by the polymerase chain reaction and detected by ethidium bromide staining after electrophoresis through agarose gel (RT-PCR). Of 217 specimens tested, 106 were positive by antigen detection, 99 by RT-PCR, and 92 by virus isolation. In a series of 97 sequential NPS specimens from 15 infants in whom RS virus induced bronchiolitis was confirmed, antigen detection proved most sensitive in the first week after onset and RT-PCR detected most positive specimens in the second week. Storage of the cells as a pellet proved more satisfactory than storage as a suspension. A further round of amplification using nested primers increased the number of positive results obtained by RT-PCR. The sensitivity of antigen detection using directly labelled monoclonal antibody to RS virus was slightly higher than that of RT-PCR, but the specificity was slightly lower.
直接检测鼻咽分泌物（NPS）中的呼吸道合胞（RS）病毒抗原可提供针对RS感染的最快速诊断测试，但在病毒脱落研究中，更灵敏的方法可能更具优势。RS病毒RNA从储存在-70℃的细胞中提取，细胞要么悬浮于添加了RNA酶抑制剂的溶液中，要么以无抑制剂的沉淀形式保存。RNA经逆转录，所得的cDNA通过聚合酶链反应进行扩增，并在通过琼脂糖凝胶电泳后用溴化乙锭染色进行检测（逆转录聚合酶链反应，RT-PCR）。在检测的217份标本中，106份抗原检测呈阳性，99份RT-PCR检测呈阳性，92份病毒分离呈阳性。在一系列来自15名确诊为RS病毒诱发细支气管炎的婴儿的97份连续NPS标本中，抗原检测在发病后第一周最为灵敏，而RT-PCR在第二周检测到的阳性标本最多。将细胞保存为沉淀比保存为悬浮液更令人满意。使用巢式引物进行的第二轮扩增增加了RT-PCR获得的阳性结果数量。使用直接标记的抗RS病毒单克隆抗体进行抗原检测的灵敏度略高于RT-PCR，但特异性略低。
