Ushijima H, Koike H, Mukoyama A, Hasegawa A, Nishimura S, Gentsch J
National Institute of Health, Tokyo, Japan.
J Med Virol. 1992 Dec;38(4):292-7. doi: 10.1002/jmv.1890380412.
Direct rotavirus serotyping (VP7, G type) in stool specimens was carried out by reverse transcription and polymerase chain reaction amplification (RT-PCR) and compared to serotyping by enzyme immunoassay with monoclonal antibodies (EIA-MAb). The methods used for double-stranded (ds) RNA extraction, RT-PCR amplification, and the primers used were modified from previous reports [Gouvea et al.: Journal of Clinical Microbiology 29:519-523, 1990; Gentsch et al.: Journal of Clinical Microbiology, 1992]. For samples that were positive by both methods, the serotypes obtained were identical, however RT-PCR typing was found to be considerably more sensitive (70.4% samples serotyped) than EIA-MAb (35.6% of samples serotyped). The overall sensitivities for detection of rotavirus in stool samples by latex agglutination, enzyme immunoassay, electron microscopy, polyacrylamide gel electrophoresis, and RT-PCR were essentially the same. The results confirm that RT-PCR typing (genotyping) is extremely valuable for G typing of samples which cannot be typed by EIA-MAb. We also developed a PCR confirmation technique for serotypes 1, 2, and 4.
通过逆转录和聚合酶链反应扩增（RT-PCR）对粪便标本进行轮状病毒直接血清分型（VP7，G型），并与使用单克隆抗体的酶免疫测定法（EIA-MAb）进行血清分型相比较。用于双链（ds）RNA提取、RT-PCR扩增的方法以及所使用的引物均根据先前的报告进行了修改[古韦亚等人：《临床微生物学杂志》29:519 - 523，1990年；根茨等人：《临床微生物学杂志》，1992年]。对于两种方法均呈阳性的样本，所获得的血清型相同，然而发现RT-PCR分型比EIA-MAb分型的灵敏度要高得多（70.4%的样本可进行血清分型），而EIA-MAb分型的样本比例为35.6%。通过乳胶凝集试验、酶免疫测定法、电子显微镜检查、聚丙烯酰胺凝胶电泳和RT-PCR检测粪便样本中轮状病毒的总体灵敏度基本相同。结果证实，RT-PCR分型（基因分型）对于无法通过EIA-MAb进行分型的样本的G分型极具价值。我们还开发了针对1、2和4型血清型的PCR确认技术。
