Detection of Human cytomegalovirus Infection by PCR-Hybridization-ELISA

YAN Ying; CHEN xiao; YAN yuan; DONG Chang

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April 2003

YAN Ying, CHEN xiao, YAN yuan and DONG Chang. Detection of Human cytomegalovirus Infection by PCR-Hybridization-ELISA[J]. Virologica Sinica, 2003, 18(2): 115-118.

Citation: YAN Ying, CHEN xiao, YAN yuan, DONG Chang. Detection of Human cytomegalovirus Infection by PCR-Hybridization-ELISA .VIROLOGICA SINICA, 2003, 18(2) : 115-118.

PCR一微孔板杂交一ELISA法检测巨细胞病毒感染

1.
1．武汉大学医学院病毒研究所，湖北武汉，430071：2．武汉市铁路第一中学，湖北武汉，430033

摘要

建立PCR．微孑L板杂交一ELISA法检测人血清标本中巨细胞病毒DNA。将5’端标记生物素的PCR扩增产物与5’端标记地高辛的探针呈液相混合，90~C 2min，55"C，lmin杂交。杂交后产物被链霉亲和素酶标板固定，经酶标记抗地高辛标记抗体结合显色。本试剂检测灵敏度为2．5×104 copies／mL，比传统PCR和ELISA敏度性高，特异性强，与单纯疱疹病毒I型和II型，风疹病毒、EB病毒、腺病毒核酸无交叉反应，批内CV为8．9％，批间 CV为10．5％。该法可用于定性和定量检测HCMV DNA。
- 聚合酶链反应
- , 酶联免疫反应
- , 巨细胞病毒

Detection of Human cytomegalovirus Infection by PCR-Hybridization-ELISA

1.
The Virus Institute of Medical College of Wuhan University,Wuhan 430071，China，2．The First Railway Meddle School,Wuhan 430033，China

Abstract

A PCR—M PH—ELISA method detecting HCMV infection was established．The PCR products of HCMV DNA were binting—labeled by a 5’-biotinlabeled primer through sample am plification．After the labeled product was mixed with a 5’-digoxin—labeled probe，hybridization was carried out for 2 mi nutes at 90℃ and for 1 minute at 55℃ ．then the hybridized product was captured on microplate wells coated with streptoavidin and detected with a peroxidase—labeled an tibody and TMB substrate． The sensitivity of the method was 2．5×1 04 copies／mL ． and higher than that of conventiona1 PCR and ELISA method．The results did not cross—react with that of HSV1，HSV2，RuV,EBV and ADV DNA． Th e method has potentia1 use for qualitative an d quantitative detection of HCMV infection．
- PCR
- , ELISA
- , Human cytomegaloirus

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Detection of Human cytomegalovirus Infection by PCR-Hybridization-ELISA

1. The Virus Institute of Medical College of Wuhan University,Wuhan 430071，China，2．The First Railway Meddle School,Wuhan 430033，China

Abstract: A PCR—M PH—ELISA method detecting HCMV infection was established．The PCR products of HCMV DNA were binting—labeled by a 5’-biotinlabeled primer through sample am plification．After the labeled product was mixed with a 5’-digoxin—labeled probe，hybridization was carried out for 2 mi nutes at 90℃ and for 1 minute at 55℃ ．then the hybridized product was captured on microplate wells coated with streptoavidin and detected with a peroxidase—labeled an tibody and TMB substrate． The sensitivity of the method was 2．5×1 04 copies／mL ． and higher than that of conventiona1 PCR and ELISA method．The results did not cross—react with that of HSV1，HSV2，RuV,EBV and ADV DNA． Th e method has potentia1 use for qualitative an d quantitative detection of HCMV infection．