Closed-tube saltatory rolling circle amplification with hydroxynaphthol blue for visual on-site detection of peanut as an allergenic food

Peanut allergy as one of the major causes of food-induced anaphylaxis is usually life-threatening for specific individuals. Nowadays, the effective remedy is avoidance of consuming allergen-containing food, urging monitoring the presence of peanut in foods. Herein, we proposed a closed-tube saltatory rolling circle amplification (SRCA) assay with hydroxynaphthol blue (HNB) for visual on-site detection of peanut allergen, displaying highly specificity for Ara h 1-encoding gene. The results can be discriminated by naked eye without opening the tube, averting costly sophisticated apparatus and reducing the risk of cross-contamination. The assay was 1000-, 100- and 10-fold more sensitive than PCR, LAMP and SRCA, respectively. The relative limit of detection is 0.01% in a binary mixture. The relative sensitivity, specificity and accuracy of this method were 100%, 97.50% and 98.67%, respectively. Thus, the visual closed-tube HNB-SRCA assay with superiorities of easy-to-perform, cost-effective and user-friendly has excellent potential for on-site application of food allergy.