Development Of New Immunoassay Methods For The Detection Of Gentamicin

Gentamicin is an aminoglycoside antibiotic which was widely used in treating bacterial infections in animals because of its good spectrum of activity against gram-positive and gram-negative bacteria. The presence of gentamicin residues in food is considered as a high risk to consumer. The objective of our research is to develop a rapid and reliable analytical method, which is used to monitor gentamicin residue in animal tissues. The immunogen and the coating antigen were synthesized by using gentamicin coupled to carrier protein bovine serum albumin (BSA) and ovalbumin (OVA) respectively. After the polyclonal antibodies against gentamicin were obtained, an ELISA method for the determination of Gentamicin was developed by using the polyclonal antibodies.The antiserum against gentamicin was acquired after immunization in rabbits.The titer of antiserum was 512000. To develop gentamicin immunoassay, the influence of several factors such as coating time, competition time, and pH were studied. Under optimized conditions, the detection limit was 0.1ng/ml. The results indicated that the optimal working dilution of coating antigen and antibody was 1:3200 and 1:16000. The best concentrtion of enzyme labled secondary antibody was 1:4000. To test the speciality of antibody, three aminoglycoside drugs were used in the competition ELISA as the competition antigen.The results indicated that the antibody no cross-reaction with other aminoglycodides.By using the antibody,We establihed immunochromatographic strip for gentamicin. The detection limit of this assay is 50ng/ml,which is sensitive enough to screen gentamicin residual .Acording to real sample analysis,the result agrees well with Enzyme-linked immunosorbent assay(ELISA).We also synthesized magnetic resonance imaging immuinosensor for the detection of gentamicin .We successfully established the standard curve of gentamicin with a equations: y=1.775x-2.44.The detection range for gentamicin is 0.91-24ng/ml.Thus this dection method has a fully advantage against the other methods .