title: [The establishment of a novel method of nano-immunomagnetic separation and Real-time PCR for detecting Vibrio cholerae from seafood].
abstract: OBJECTIVE
  A novel method of Nano-Immunomagnetic Separation (Nano-IMS) plus Real-time PCR was established for detecting Vibrio cholerae.
  
  
  METHODS
  The Nano-Immunomagnetic Beads were created by using the monoclonal antibody of Vibrio cholerae, which was named Nano-IMB-Vc. Nano-IMB-Vc has specific adsorption of Vibrio cholerae, combined with Real-time PCR technology, a method for rapid detection of Vibrio cholerae was established. The capture specificity of Nano-IMB-Vc was tested by using 15 bacteria strains. The specificity of Real-time PCR method was tested by using 102 targets and 101 non-targets bacteria strains. The sensitivity of Nano-IMS plus Real-time PCR were tested in pure culture and in artificial samples and compared with NMKL No.156.
  
  
  RESULTS
  The capture ratio of Nano-IMB-Vc was reached 70.2% at the level of 10(3) CFU/ml. In pure culture, the sensitivity of Nano-IMS plus Real-time PCR was reached at 5.4×10(2) CFU/ml. The specific of Real-time PCR method was tested by using 102 targets and 101 non-targets bacteria. The results showed that 102 strains of Vibrio cholerae test results were all positive, and the rest of the 101 strains of non-target bacteria test results were negative. No cross-reaction was founded. Add 1 CFU vibrio cholerae per 25 g sample, it could be detect with Nano-IMS plus Real-time PCR method after 8 hours enrichment.
  
  
  CONCLUSIONS
  The Nano-IMS plus Real-time PCR method of Vibrio cholerae established in this study has good specificity and sensitivity, which could be applied to the rapid detection of Vibrio cholerae.
authors.affiliation: College of Food Science and Engineering, Beijing University of Agriculture, Beijing 102206, China.
authors.forename: Jinxia
authors.initials: J
authors.lastname: Cheng
authors.affiliation: Email: zengj@bjciq.gov.cn.
authors.forename: Jing
authors.initials: J
authors.lastname: Zeng
authors.forename: Li
authors.initials: L
authors.lastname: Liu
authors.forename: Haiyan
authors.initials: H
authors.lastname: Wei
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Zhao
authors.forename: Ximeng
authors.initials: X
authors.lastname: Zhang
authors.forename: Lei
authors.initials: L
authors.lastname: Zhang
authors.forename: Haiyu
authors.initials: H
authors.lastname: Zhang
mesh_terms: D005516:Food Microbiology / Q000379:methods; D018189:Immunomagnetic Separation / Q000379:methods*; D036103:Nanotechnology; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D017747:Seafood / Q000032:analysis / Q000382:microbiology; D014734:Vibrio cholerae / Q000235:genetics / Q000302:isolation & purification*; D014736:Vibrio parahaemolyticus / Q000235:genetics / Q000302:isolation & purification