Establishment of an Enzyme-Linked Immunosorbent Assay Method for Detection of Heterocyclic Amines

A broad-spectrum heterocyclic aromatic amine polyclonal antibody was prepared and an indirect competitive enzyme-linked immunosorbent assay(ic-ELISA)was developed to simultaneously detect multiple heterocyclic aromatic amines. MeIQx was applied to synthesize the hapten via reacting with monomethyl succinate chloride(MCO). The polyclonal antibody was prepared by coupling hapten to protein by activated ester method to prepare immunogen,and the ic-ELISA method was established finally. The sensitivity of this ELISA(IC₅₀,for MeIQx)was 81.16 μg/L and the limit of detection(IC₁₅)was 12.07 μg/L. This method had similar recognition capacity for the quinoline heterocyclic aromatic amines(IQ,MeIQ),quinoxaline heterocyclic aromatic amines(IQx,MeIQx,4,8-DiMeIQx,7,8-DiMeIQx,4,7,8-TriMeIQx)and pyridines(PhIP)and the cross-reaction rates reached more than 93%.The recoveries of heterocyclic aromatic amine(MeIQx)in fried beef and dried meat floss samples were between 91.18% and 98.64%. There was a good correlation(R²=0.9927)between the ELISA and liquid chromatography-tandem mass spectrometry(LC-MS/MS)for the detection of MeIQx in the spiked samples. The ELISA established in this paper can be applied to detect the total amount of quinoline,quinoxaline and pyridine heterocyclic aromatic amine,and provide a simple and accurate approach for the detection of multiple heterocyclic aromatic amines in processed meat products.