The establishment of a novel method of nano-immunomagnetic separation and Real-time PCR for detecting Vibrio cholerae from seafood.
- Author:
Jinxia CHENG
1
;
Jing ZENG
2
;
Li LIU
;
Haiyan WEI
;
Xiaojuan ZHAO
;
Ximeng ZHANG
;
Lei ZHANG
;
Haiyu ZHANG
Author Information
- Publication Type:Journal Article
- MeSH: Food Microbiology; methods; Immunomagnetic Separation; methods; Nanotechnology; Real-Time Polymerase Chain Reaction; methods; Seafood; analysis; microbiology; Vibrio cholerae; genetics; isolation & purification; Vibrio parahaemolyticus; genetics; isolation & purification
- From: Chinese Journal of Preventive Medicine 2014;48(2):133-137
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVEA novel method of Nano-Immunomagnetic Separation (Nano-IMS) plus Real-time PCR was established for detecting Vibrio cholerae.
METHODSThe 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.
RESULTSThe 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.
CONCLUSIONSThe 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.
