Establishment and evaluation of fluorescence isothermal amplification assay for the detection of Enterovirus 71

doi:10.3969/j.issn.1673-8640.2014.11.009

Abstract

Abstract:

Objective　To establish a rapid and reliable fluorescence isothermal amplification assay (SAT) for the detection of Enterovirus 71 (EV71). Methods　By designing specific primers and optimal probe and using M-MLV reverse transcriptase and T7 RNA polymerase, the SAT was developed for detecting EV71 based on isothermal RNA amplification and fluorescence quantitation polymerase chain reaction (PCR). A total of 199 stool specimens from the children with hand, foot, mouth disease from Children's Hospital of Fudan University were determined. Using EV71 PCR fluorescence diagnostic kits as reference kit and DNA sequencing as the third party verification method, the Kappa analysis was performed on the study data for consistency. Results　A total of 119 stool specimens were positive by SAT, and there were 21 specimens being negative by PCR-fluorescence probe assay, but 20 specimens were confirmed to be positive by DNA sequencing. The Kappa value between SAT and PCR-fluorescence probe assay was 0.789. The sensitivity and specificity of SAT were 100% and 98.77%, respectively. Conclusions　This study indicates that RNA SAT for the detection of EV71 is a sensitive, specific, accurate, rapid and reliable method, and it should be a potential method for the rapid detection of EV71 infection.

Key words: Enterovirus 71, Fluorescence isothermal amplification assay, Hand, foot, mouth disease, Sensitivity, Specificity

CLC Number:

R446.1

CAO Lingfeng, SU Liyun, DONG Niuniu, XU Jin.. Establishment and evaluation of fluorescence isothermal amplification assay for the detection of Enterovirus 71[J]. , 2014, 29(11): 1115-1119.

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