Preparation of Artificial Antigen and Development of IgY-Based Indirect Competitive ELISA for the Detection of Kanamycin Residues

The present study was aimed to evaluate the feasibility to use chicken egg yolk antibodies (IgY) for the detection of Kanamycin (Kana) residues in food stuffs. Bovine serum albumin (BSA) was conjugated with Kana by carbodiimide method, and the laying chickens were immunized with Kana-BSA conjugate. PEG-6000 method was employed to extract IgY. The peak titer of anti-Kana IgY was 1:256,000 after the fifth booster immunization. The optimal dilution for anti-Kana IgY was 1:25,000 to obtain optical density (OD) of 1.0 at 2 mu g/mL of OVA-Kana coating concentration. The indirect competitive ELISA (ic-ELISA) showed that the IC50 value of anti-Kana IgY was 4.48 ng/mL and the regression curve equation was y = -15.62x + 60.17 (R (2) = 0.98, n = 3). The ic-ELISA showed a lower cross-reactivity (less than 0.01 %) with other drugs (except gentamicin-1.91 %). Recoveries from milk, pork, and chicken powder samples were in the range of 82.02 to 98.20 %, with relative standard deviation lower than 5.13 %. Our results indicate that generated anti-Kana IgY can be used in routine screening analysis of Kana residues in food samples.