A novel visual-mixed-dye for LAMP and its application in the detection of foodborne pathogens

Loop-mediated isothermal amplification (LAMP) has been widely applied for the detection of foodborne pathogens. Obtaining a simple, accurate readout of LAMP reaction results is crucial. Herein, a visual-mixed-dye (VMD) containing calcein (precombined with MnCl2) and hydroxynaphthol blue (HNB) for LAMP end-point detection was developed. The optimal final concentrations of these components in VMD were 25 and 300 μM, respectively. Due to the formation of HNBMn2+ during the LAMP reaction, the VMD-loaded assay exhibited superior visual properties, changing from light gray (negative) to dark blue (positive) under natural light. Additionally, compared with traditional single calcein or HNB dye, a weakly positive result with the VMD was purple, making it easier to distinguish by the naked eye. The visual sensitivity reached down to 20.9 copies/μL, which was comparable to that based on fluorescence. In food-contaminated samples, the practicality of VMD was verified by Vibrio parahaemolyticus and Staphylococcus aureus detection with excellent specificity. Moreover, the VMD was stable over a period of 3 months. Collectively, these findings establish the VMD as a novel dye for LAMP. [Display omitted] •A visual-mixed-dye (VMD) containing calcein and HNB was used for LAMP detection.•The result of VMD-loaded LAMP could be directly discriminated via naked eyes.•VMD made the color difference between negative and positive assays more obvious.•The use of VMD made the judgement of weak-positive result more clear and accurate.•The VMD-loaded assay could detect food samples contaminated by foodborne pathogens.