Synthesis of furazolidone metabolite artificial antigen and development of indirect competitive chemiluminescence enzyme immunoassay

An indirect competitive chemiluminescence enzyme immunoassay ( ic-CLEIA) was used for determination and identification of the furazolidone metabolite in animal-derived food. AOZ can be translated into hapten CPAOZ by derivative reaction.CPAOZ-OVA, as the coating antigen, was prepared by activated ester method.After optimizing the dilution multiple of coating antigen and anti-body, blocking buffer, competitive reaction time and the dilution multiple of second antibody, a standard curve was established, and finally the specificity of the method was evaluated.The results indicated that, the optimum dilution multiple of coating antigen was 2000-fold, the anti-body was 20000-fold.Blocking buffer was dried skim milk at the concentration of 2%.The competitive reaction time was 2 h, and the ideal dilution multiple of secondary antibody was 4000-fold.The linearity range was 0.075⁷.396 ng/m L, and half-inhibitory concentration ( IC₅₀) value was 0.74 ng/m L.The crossreactivity value between AOZ and furazolidone original drug was 23.4%, and cross reactivity value with other analogues and derivatives was < 0.1%. The method with demonstrated excellent sensitivity and specificity can provide the basis for rapid detection of AOZ content in animal-derived food for supervision department.