A Modified Quantum Dot-Based Dot Blot Assay for Rapid Detection of Fish Pathogen Vibrio anguillarum.
Vibrio anguillarum, a devastating pathogen causing vibriosis among marine fish, is prevailing in worldwide fishery industries and accounts for grievous economic losses. Therefore, a rapid on-site detection and diagnostic technique for this pathogen is in urgent need. In this study, two mouse monoclonal antibodies (MAbs) against V. anguillarum, 6B3-C5 and 8G3-B5, were generated by using hybridoma technology and their isotypes were characterized. MAb 6B3-C5 was chosen as the detector antibody and conjugated with quantum dots. Based on MAb 6B3- C5 labeled with quantum dots, a modified dot blot assay was developed for the on-site determination of V. anguillarum. It was found that the method had no cross-reactivity with other than V. anguillarum bacteria. The detection limit (LOD) for V. anguillarum was 1 × 10(3) CFU/ml in cultured bacterial suspension samples, which was a 100-fold higher sensitivity than the reported colloidal gold immunochromatographic test strip. When V. anguillarum was mixed with turbot tissue homogenates, the LOD was 1 × 10(3) CFU/ml, suggesting that tissue homogenates did not influence the detection capabilities. Preenrichment with the tissue homogenates for 12 h could raise the LOD up to 1 × 10(2) CFU/ml, confirming the reliability of the method.
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.|Shanghai Engineering Research Center of Maricultured Animal Vaccines, Shanghai 200237, China.
Yang
Y
Zhang
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.|Shanghai Engineering Research Center of Maricultured Animal Vaccines, Shanghai 200237, China.
Jingfan
J
Xiao
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.|Shanghai Engineering Research Center of Maricultured Animal Vaccines, Shanghai 200237, China.|Shanghai Collaborative Innovation Center for Biomanufacturing, Shanghai 200237, China.
Qiyao
Q
Wang
State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.|Shanghai Engineering Research Center of Maricultured Animal Vaccines, Shanghai 200237, China.|Shanghai Collaborative Innovation Center for Biomanufacturing, Shanghai 200237, China.
Yuanxing
Y
Zhang
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D005393:Fish Diseases / Q000175:diagnosis* / Q000382:microbiology; D005412:Flatfishes / Q000382:microbiology; D015151:Immunoblotting / Q000379:methods*; D057230:Limit of Detection; D051379:Mice; D045663:Quantum Dots; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014733:Vibrio / Q000276:immunology / Q000302:isolation & purification*; D014735:Vibrio Infections / Q000175:diagnosis* / Q000382:microbiology
D000911:Antibodies, Monoclonal
Fish pathogen; Vibrio anguillarum; quantum dot; rapid detection