An indirect competitive enzyme-linked immunosorbent assay for acrylamide detection based on a monoclonal antibody

Acrylamide (AA) is formed spontaneously in heated foodstuffs and is a focus of concern in many people due to safety. In this study, we developed an indirect competitive enzyme-linked immunosorbent assay (icELISA) based on a monoclonal antibody (MAb) to detect a derivative, which was generated from 4-mercaptobenzoic acid (4-MBA). As AA is a very small molecule (71.08 Da) and cannot elicit a homologous monoclonal antibody, we coupled the AA derivative (AA-4-MBA) to a carrier protein such as bovine serum albumin (BSA) and ovalbumin (OVA). The conjugates were used as the immunogen and coating antigen. A rapid and sensitive icELISA against AA-4-MBA was obtained by optimizing the experimental parameters. The MAb which had no specificity for AA or 4-MBA, but had high affinity for AA-4-MBA, had a satisfactory IC₅₀ of 32 ng/ml and a limit of detection of 8.87 ng/ml. The quantitative working range was 8.87–112.92 ng/ml (IC₂₀ to IC₈₀). Cross-reactivity with other analogues was lower than 10%. These results indicated that the developed icELISA was a fast and efficient method for detecting AA in food.