Development Of A Colloidal Gold Immunochromatographic Assay For Detection Of Florfenicol In Pork

In recent years, the issue about veterinary drug residues in food related animal products, such as "clenbuterol hydrochloride event", "turbot event", occurs frequently in China. Safety of animal products is concerned by more and more people. According to Chinese first national inventory about usage and emissions on antibiotics which was published in 2015, China had consumed 162,000 tons of antibiotics in 2013. About 52% of antibiotics were used in livestock and poultry breeding. The proportion of veterinary antibiotics was up to 84.3% in 36 kinds of common antibiotics.Florfenicol is used broadly in livestock and poultry breeding as it is a broad-spectrum antibiotics. It has blood toxicity, embryo toxicity and immunosuppression. It brings great harm to human when eating much pork contained excessive florfenicol residues. At the same time, abusing florfenicol can enhance bacterial antibiotics resistance and has an adverse impact on our environment and economy. In this study, a method for rapid detection of florfenicol residues in pork was established.The muscles and livers of pig from Nanchang City, Jiujiang City, Fuzhou City and Yichun City were screened. The result showed the antibiotics was used under controlled. However several antibiotics such as florfeniol and enrofloxacin had high positive rate.Florfenicol amine(FFA) was conjugated to carrier protein by glutaraldehyde method and carbodiimide method. The conjugates were characterized by ultraviolet-visible spctrophotometer. Then the holoantigen prepared and holoantigen bought from the company were used to immune mice. The highest titer of antiserum is 1:32000.A method for rapid detection of florfenicol residues in pork was established by the experiments of selecting antigen and antibody, adjusting pH of the labeling process, optimizing the concentration of antigen and antibody sprayed on T and C lines and determining reaction time. The optimum conditions were as follows: the optimum pH of labeling process was 8.0, the labeling mass ratio was 6 ?g antibody per milliliter colloidal gold, the amount of gold-labelled antibody was 5 ?L, the concentration of antigen and antibody sprayed on T and C line was 1.0 mg/m L and 0.3 mg/m L, respectively, the detection time was 25 min. The assay had a liner range from 4~200 ?g/L. The IC50 was 9.795 ?g/L and the limit of detection was 2.18 ?g/L. The intra assay recoveries ranged from 99~113% and coefficient of variation(CV) ranged from 10.1~15.6%. The inter assay recoveries ranged from 98.4~118.3% and CV ranged from 14.3~17.2%. This method can be applied to rapid quantitative detection of florfenicol in pork with a good correlation to commercial ELISA kits.