东南大学学报(医学版)
東南大學學報(醫學版)
동남대학학보(의학판)
JOURNAL OF SOUTHEAST UNIVERSITY(MEDICAL SCIENCE EDITION)
2013年
6期
698-703
,共6页
吴菲菲%董敏%葛路遥%胡馨云%张程%张建华%江炳福%孟继鸿
吳菲菲%董敏%葛路遙%鬍馨雲%張程%張建華%江炳福%孟繼鴻
오비비%동민%갈로요%호형운%장정%장건화%강병복%맹계홍
手足口病%肠道病毒71型%单克隆抗体%免疫荧光法%病毒分离
手足口病%腸道病毒71型%單剋隆抗體%免疫熒光法%病毒分離
수족구병%장도병독71형%단극륭항체%면역형광법%병독분리
hand,foot and mouth disease%enterovirus 71%monoclonal antibody%immunofluorescence assay%virus isolation
目的:建立肠道病毒71型( EV71)特异性检测的免疫荧光方法用于病毒分离培养后的EV71鉴定,为EV71生物学特征研究和疫苗株的筛选奠定基础。方法:以EV71 VP1为免疫原制备单克隆抗体,用经测序鉴定的肠道病毒毒株建立EV71特异性免疫荧光方法;将不同地区收集的手足口病患儿的临床标本进行病毒分离,利用EV71特异性单抗进行分离培养后的病毒鉴定。结果:制备获得2株单抗,经鉴定其中1株3 E12为EV71特异性单抗;以此单抗建立的免疫荧光方法对EV71感染细胞呈阳性反应,而与柯萨基病毒A16和埃可病毒6等非EV71肠道病毒不反应。从104份手足口病临床标本中共分离到35株病毒,产生肠道病毒典型的细胞病变;用3E12特异性单抗免疫荧光鉴定分离病毒,其中29株为EV71,与RT-PCR验证结果完全一致。结论:成功制备获得EV71特异性单克隆抗体,以此单抗建立的EV71特异性免疫荧光方法敏感性高、特异性强,可用于分离培养EV71的鉴定。
目的:建立腸道病毒71型( EV71)特異性檢測的免疫熒光方法用于病毒分離培養後的EV71鑒定,為EV71生物學特徵研究和疫苗株的篩選奠定基礎。方法:以EV71 VP1為免疫原製備單剋隆抗體,用經測序鑒定的腸道病毒毒株建立EV71特異性免疫熒光方法;將不同地區收集的手足口病患兒的臨床標本進行病毒分離,利用EV71特異性單抗進行分離培養後的病毒鑒定。結果:製備穫得2株單抗,經鑒定其中1株3 E12為EV71特異性單抗;以此單抗建立的免疫熒光方法對EV71感染細胞呈暘性反應,而與柯薩基病毒A16和埃可病毒6等非EV71腸道病毒不反應。從104份手足口病臨床標本中共分離到35株病毒,產生腸道病毒典型的細胞病變;用3E12特異性單抗免疫熒光鑒定分離病毒,其中29株為EV71,與RT-PCR驗證結果完全一緻。結論:成功製備穫得EV71特異性單剋隆抗體,以此單抗建立的EV71特異性免疫熒光方法敏感性高、特異性彊,可用于分離培養EV71的鑒定。
목적:건립장도병독71형( EV71)특이성검측적면역형광방법용우병독분리배양후적EV71감정,위EV71생물학특정연구화역묘주적사선전정기출。방법:이EV71 VP1위면역원제비단극륭항체,용경측서감정적장도병독독주건립EV71특이성면역형광방법;장불동지구수집적수족구병환인적림상표본진행병독분리,이용EV71특이성단항진행분리배양후적병독감정。결과:제비획득2주단항,경감정기중1주3 E12위EV71특이성단항;이차단항건립적면역형광방법대EV71감염세포정양성반응,이여가살기병독A16화애가병독6등비EV71장도병독불반응。종104빈수족구병림상표본중공분리도35주병독,산생장도병독전형적세포병변;용3E12특이성단항면역형광감정분리병독,기중29주위EV71,여RT-PCR험증결과완전일치。결론:성공제비획득EV71특이성단극륭항체,이차단항건립적EV71특이성면역형광방법민감성고、특이성강,가용우분리배양EV71적감정。
Objective: To establish a specific immunofluorescence assay for identification of enterovirus 71 ( EV71 ) after isolation from cell culture and lay the foundation for EV 71 pathogen biology research and vaccine strains screening.Methods:Monoclonal antibodies (McAb) were prepared against EV71 VP1.An EV71-specific McAb and identified enteroviruses were used for establishment of an EV 71-specific immunoflurescence assay .The assay was applied for EV71 identification after virus isolation from clinical samples collected from patients with hand, foot and mouth disease ( HFMD) at different areas of China .Results: Two McAbs, designated 3E12 and 5C1, against EV71 VP1 were obtained.The McAb 3E12 was EV71-specific.The immunofluorescence assay established based on 3E12 was identified to detect EV71 only.Other enterovirus strains such as coxsackievirus A 16 and ECHO virus 6 were not detected by the assay .Thirty-five virus strains were isolated with typical cytopathogenic effect of enterovirus in cell culture from 104 HFMD specimens .Twenty-nine isolated virus strains were identified as EV71 by the EV71-specific immunofluorescence assay , which was the same as the result tested by EV 71 RT-PCR. Conclusion: The EV71-specific McAb is successfully prepared and used for establishment of the immunofluorescence assay with high sensitivity and specificity for identification of EV 71 isolated from cell culture .