title: [Development of EV71, CA16 and other enterovirus vrial real-time qualitative PCR diagnostic kit].
abstract: OBJECTIVE
  A novel multiplex real-time RT-PCR kit was developed to detect EV71, CoxA16 and other human enteroviruses simultaneously with an internal amplification control to avoids false negatives, which used for hand, foot and mouth disease in the clinical diagnosis and epidemiological surveillance.
  
  
  METHODS
  Design specific primers and probes of EV71, CA16, other intestinal virus and internal amplification control, improve the extraction method of virus nucleic acid. Optimization the detection system of real-time quantitative PCR. Research the products of the accuracy, stability, precision, amplification efficiency and detection of linear range.
  
  
  RESULTS
  The primers and probes had high spicificity. The Viral RNA extraction effect of this Kit is as same as that of QIAamp Viral RNA mini Kit (QIAGEN company), but less reagent cost. The optimal concentrations of primers and probes are 0.2 micromol/L for all the upstream and downstream primers, 0.06 micromol/L for probes of other human enteroviruse, 0.08 micromol/L for probes of EV71 and CA16 respectively. The kit has good stability, accuracy and precision. The amplification efficiencies of EV71, CoxA16 and other human enteroviruses are 106% ,101% and 105% and the detection of linear range is from 10(9) copies/microl-10(2) copies/microl.
  
  
  CONCLUSIONS
  The novel multiplex real-time RT-PCR kit for detecting EV71, CoxA16 and other human enteroviruses simultaneously with an internal amplification control has good stability, accuracy, precision and amplification efficiencies. So it has great value in clinical application.
authors: Huzhou Central Hospital, Zhejiang 313000, China.
authors: Li-Qin
authors: LQ
authors: Li
authors: Jing
authors: J
authors: Zhong
authors: Lin-Fu
authors: LF
authors: Zhou
authors: Fu-Chu
authors: FC
authors: Qian
authors: Jia-Wei
authors: JW
authors: Wang
authors: Li-Cheng
authors: LC
authors: Dai
mesh_terms: D004770:Enterovirus / Q000302:isolation & purification*; D006801:Humans; D011933:Reagent Kits, Diagnostic; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*
chemical_list: D011933:Reagent Kits, Diagnostic