Abstract: In this study, a rapid, accurate and stable direct competitive ELISA (dcELISA) for the detection of chloramphenicol residue in honey was developed. The anti-chloramphenicol monoclonal antibody was used as coating antigen, chloramphenicol-horseradish peroxidase conjugate as marker and 3,3’,5,5’-tetramethyl benzidine (TMB) as chromogenic substrate. Six main experimental conditions, including buffer for chloramphenicol standard, coating temperature, and dilution of coating antibody were optimized. In addition, the accuracy and stability of the method were also investigated. The obtained standard curve equation was y = ?17.425x + 97.509 (R2 = 0.991 2), and the IC50 was 0.63 ng/mL. The limit of detection (LOD) and linear rang were (0.04 ± 0.01) ng/mL and 0.10?4.17 ng/mL, respectively. The whole detection process required about 40 min. In honey samples, the LOD, LOQ and spiked recovery were 0.15 ng/g, 0.33 ng/g and 97.58%?100.94%, respectively. The intra-batch and inter-batch coefficients of variation were both less than 11%. These results indicated that the developed dcELISA method possessed high accuracy and stability.

Key words: chloramphenicol, honey, direct competitive ELISA, optimization