Combination of loop-mediated isothermal amplification and AuNP-oligoprobe colourimetric assay for pork authentication in processed-meat products

Abstract

Pork adulteration is a major concern for Muslims and Jews whose diets are restricted by religious beliefs, as well as those who are allergic to pork meat and its derivatives. Accurate pork authentication is of great importance to assist this demographic group of people in making decision on their product purchase. The aim of this study was to develop a new analytical method for pork authentication in processed-meat products based on a combination of loop-mediated isothermal amplification (LAMP) and AuNP-nanoprobe colourimetric assay. The LAMP conditions were first optimised to obtain the highest yield of amplified DNA products within the shortest time. Oligoprobe-functionalised AuNPs were then hybridised with LAMP-DNA amplicons, and subsequently challenged with MgSO4 at a high concentration to induce AuNP aggregation. In the presence of pork DNA, the colloidal AuNPs-probe remained unchanged in its red colour, which indicates the dispersion of AuNPs. In contrast, in the absence of pork DNA, the colour was changed to colourless as a result from the aggregation of AuNPs. The LAMP-AuNP-nanoprobe assay offers a high sensitivity with a limit of detection as low as 100 pg of pork DNA. The assay is highly specific to pork content without cross-reactivity with the other meat species tested. The assay developed herein can become a simple, inexpensive, precise, and rapid analytical tool for small laboratories or the general public interested in halal food authentication.