Development of an indirect ELISA for Acid Red 73 in Shrimps

Acid red 73( AR73) was coupled to bovine serum albumin( BSA) and ovalbumin( OVA) by the use of N- hydroxysuccinimide active method and the CDI method to prepare the immunogen and coating antigen,respectively.The anti- AR73 antibody were obtained and purified by the caprylic acid- ammonium sulfate method.Based on the Ig G,an indirect competitive ELISA method and optimum reaction conditions for detection of AR73 was developed.The half of inhibition concentration( IC50) was 45.39 μg /L,the limit of detection for acid red 73 was5.64 μg / L,and the cross- reactivity study showed that the polyclonal antiserum were highly specific to AR73,and no cross- reactivity was detected between the obtained polyclonal antiserum and the other competitors( Congo Red,Phenol Red,Safranine,Basic Fuchsin and Carmine) except to Sudan red Ⅲ( 0.15%).The recoveries from the standards fortified blank samples were in the range of 71.3% ⁸4.4% with Coefficient of Variance lower than 9.55%.This developed method can be used to determine the acid red 73 residue in shrimps,and can help to prepare the monoclonal antibody and develop the rapid test kits for acid red 73.