Development Of Rapid Assay For Aminoglycoside Antibiotics And Lincomycin

This study synthesized immunogens for gentamicin, kanamycin, streptomycin, neomycin by glutaraldehyde; lincomycin derivate synthesized with succinic anhydride was linked to carrier according to carbodiimide method as immunogen for lincomycin. Carbodiimide method was used to prepare the coating antigen for aminoglycoside antibiotics; EDC, DCC, MA, Na IO4 and CDI five methods were used to prepare the coating antigen for lincomycin to optimize the mice antiserum, and Na IO4 method was found the best one showing the best inhibition in ELISA. Five cell lines of monoclonal antibody were prepared and they were GEN(2F7), KAN(1E10), STR(1G12), NEO(3B5), LIN(5F6).The IC50 of gentamicin m Ab was 0.087 ng/m L; the IC50 of kanamycin m Ab was 0.07 ng/m L, and that of tobramycin was 0.054 ng/m L; the IC50 of streptomycin m Ab was 0.41 ng/m L, and that of dihydrostreptomycin was 0.34 ng/m L; the IC50 of neomycin m Ab was 0.46 ng/m L; the IC50 of lincomycin m Ab was 0.3 ng/m L, showed 0.6% cross-reactivity with clindamycin. Indirect ELISA was established to detect gentamicin, kanamycin, tobramycin, streptomycin, dihydrostreptomycin, neomycin and lincomycin in spiked milk based on the monoclonal antibody, and the limit of detection were respectively 0.45, 0.39, 0.3, 1.5, 1.24, 2.7 and 2.1 ng/m L.Based on the monoclonal antibody, colloidal gold immunochromatography assay was developed. The cut-off value of Gentamicin was 5 ng/m L. The cut-off value of kanamycin and tobramycin was 5 ng/m L. The cut-off value of streptomycin and dihydrostreptomycin was 12 ng/m L. The cut-off value of neomycin was 15 ng/m L. The cut-off value of lincomycin was 2.5 ng/m L. Then multiple strip was developed to detect five antibiotics in milk at the same time. The cutoff value of gentamycin, kanamycin and streptomycin were 50 ng/m L, that of neomycin was 100 ng/m L, and 25 ng/m L for lincomycin.