Development Of An Colloidal Gold Immunochromatographic Strip For Detection Of Streptomycin Residue In Food Derived From Animals

Streptomycin(SM) is an Aminoglycosides antibiotics.It is effective to treat cow mastitis, infectious coryza and endometritis, and is frequently added into feeds for the prevention or treatments of animal diseases. However, illegal administration will lead to residues in edible tissues, and finally do latent harm to human health, resulting in hearing toxicity and kidney toxicity. Many countries and international organizations have established the maximum residue limit of Streptomycin in animal tissues for human consumption.Many chemical and physics methods for detecting streptomycin residued in edible tissues of animals have been established. Though some of these methods are sensitive and accurate, they are not suitable for screening a large number of samples because these methods are time-consuming, costly and highly-skilled. Enzyme-linked immunosorbent assay (ELISA) and Gold Immunochromatographic assay (GICA) as two rapid, special, sensitive and safe biological methods, are gradually applied in drug residues assay.To find and establish the best way for detecting SM residues, this research synthesized the antigen, developed the antibodies and colloidal gold to establish the GICA method for detecting SM by the technology of monoclonal antibody and GICA, and finally prepared the strip.Streptomycin was conjugated with ovalbumin (OVA) separatedly with carboxymethoxylamine (CMO) method to form artifical antigen. The conjugant was identified by ELISA and UV spectrum methods. The hybridoma cell was expanded cultured and inoculated into mice abdominal cavity to produce ascites. The ascites of the mice was collected and puried to obtain the monoclonal antibody. The McAb was determined with coating antigen and the results showed that the concentration of the McAb was 4.2mg/ml, the titer of the McAb was 1:160000 and the 50% inhibition concentration was 93.22ng/ml. The cross-reactive rates of McAb to other seven antibiotics were negligible. Colloidal gold, whose diameter is about 18nm, is obtained by reducing the gold chloride with sodium citrate, and labels SM McAb. The optimum pH for labelling is about 8.5, and the amount of McAb is 108μg/ml. The glass fibre of Ahlstrom 8964 and nitrocellulose (NC) of Millipore 135 are used as chromatographic materials, in which the antigen is coated at 0.2mg/ml, the sheep anti-mouse IgG is coated at 50 times dilution, and the colloidal gold-labelled antibody is coated at 30% concentration.The major advantages of the strip were that results could be obtained within 5 min and that all needed reagents were included in the strip. The strip could be used to detect the Streptomycin residue in food derived from animals. The method of producing the one step strip for SM could be used for reference in the development of strips for the detection of other veterinary drugs.