Rapid DNA extraction and colorimetric amplicon visualisation speed up LAMP-based detection of soybean allergen in foods

Abstract

AbstractDetection of allergens in foods, including soybean, is relevant for food labelling requirements. Moreover, allergen-specific methods may allow standardisation of allergens in food matrices for use in food challenges as allergy diagnostic approaches. Rapid methods are preferred for screening and along the manufacturing line. Previously, we demonstrated sensitive and specific detection of soybean DNA by combining loop-mediated isothermal amplification (LAMP) and lateral flow device (LFD)-like visualisation. However, lengthy DNA extraction and potential contamination of subsequent by previous LAMP reactions from unclosed LFD may impact its use as a rapid and robust method. Here, we developed a rapid protocol for DNA extraction. Moreover, we identified phenol red for distinct visualisation of positive reactions in permanently closed reaction tubes. The optimised method was validated using complex foods (boiled sausage, instant soup, and chocolate) with known amounts of soybean. Further, its applicability was shown in 12 processed retail foods. Results were verified by orthogonal qPCR. The enhanced LAMP method allowed detection at or below 10 mg soybean per kg processed food. The method provides rapid and easy-to-use screening without the need for detection equipment. Hence, it may serve to verify the presence of soybean ingredients and support a risk-based precautionary labelling of non-ingredient soybean in compound foods. Also, as determination of clinical reaction thresholds before and after allergen immunotherapy (AIT) is both inclusion and exclusion criterion for clinical trials and success parameter of AIT, the method may allow verification of calculable soybean content in provocation meals and thus a standardised administration for threshold determination before and after AIT.