Double antibodies sandwich enzyme-linked immunosorbent assay for the detection of Alicyclobacillus acidoterrestris in apple juice concentrate

Two heterologous antibodies against Alicyclobacillus acidoterrestris (A. acidoterrestris) were obtained from Sprague Dawley (SD) Rats and Japan White Rabbits by immunizing them with an A. acidoterrestris strain isolated from apple juice concentrate (AJC). By using these two antibodies, we established a double antibodies sandwich ELISA (DAS-ELISA) method for the rapid detection of A. acidoterrestris in AJC, with the detection limit of 5 × 103 CFU/mL. There was no cross-reaction with seven common foodborne microorganisms by employing this established essay, and the coefficient of variation (CV) within and among independent runs were less than 5.0% and 10% respectively, indicating of satisfactory specificity and repeatability. This DAS-ELISA assay was successfully applied to detect A. acidoterrestris in AJC samples when coupled with selective incubation for 12 h, and the results were agreed well with those obtained from the routine K medium method. Further, this method shows advantage over indirect ELISA (ID-ELISA) and SPA-ELISA assays previously reported by our group, facilitating the use of this newly developed assay method in food products.