Species-specific TM-LAMP and Trident-like lateral flow biosensor for on-site authenticity detection of horse and donkey meat

Global food research is placing increasing emphasis on the on- site authenticity detection of meat since it is vital for this process to be conducted accurately and efficiently. In this study, a trident-like lateral flow biosensor (LFB) for multiple on-site authenticity detection of horse and donkey meat was established for the first time based on species-specific TM-LAMP (tag-labeled multiplex loop-mediated isothermal amplification). Firstly, a novel, accurate model for screening chromosomal endogenous reference genes was constructed using Perl as the bioinformatics language, based on which the single-copy genes LOC106782588 and LOC106825524 were selected as the endogenous reference genes for horses and donkeys, respectively. Secondly, antigen tag-labeled multiplex LAMP based lateral flow biosensor was established. Following optimization, the limit of detection (LOD) reached a level as low as 40 pg, which was equivalent to 15 copies in horses and donkeys. TM-LAMP was proved to be suitable for on-site detection since the process took only 40 min, with no need of any precision instruments. The analysis of various processed foods and mixed meat products indicated that this biosensor displayed high specificity and sensitivity consistently. As a universal platform, this biosensor can be custom-designed for on-site detection of other species by changing specific primers, which might be an excellent prospect for the identification of species authenticity.