Document Type: Full paper (Original article)
Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming 650500, PR China
Background: Salmonella can cause serious human gastroenteritis and frequently isolated from various food samples. The cell culturing, immunoassay, and polymerase chain reactions (PCRs) are the current methods to detect such pathogenic agents. However, these methods are time-consuming and labor-intensive, thus unavailable for rapid-monitoring of Salmonella. Aims: This study was to develop an immunocapture-loop-mediated isothermal amplification for rapid and sensitive detection of Salmonella. Methods: We used Salmonella as antigen to produce monoclonal antibody and mAbs were prepared via three times subcloning. The mAb 1B12 with high affinity was coated on the surface of the immuno-magnetic beads (IMBs) to capture Salmonella. The enriched products were used for loop-mediated isothermal amplification (LAMP) using the special primers targeted the conserved invA gene of Salmonella. Results: The IC-LAMP was developed based on mAb 1B12 and LAMP. Targeting the conserved invA gene of Salmonella, the detection time was shortened to 50 minutes from three days. If the reaction contains Salmonella, the green fluorescence and the trapezoidal strip can be clearly observed. Importantly, the method combines the specificity of antibody and LAMP with a detection limit of 5 CFU/mL in artificial water and milk. The results indicate that the IC-LAMP reacts only with Salmonella and has not cross-react with other similar bacteria. Conclusion: The IC-LAMP assay developed here is a rapid, sensitive, one-step-visual method to screen for the presence of Salmonella in food samples. This method is faster than traditional PCR, LAMP, and other methods, and can be used as a primary screening method for the detection.