| Aleutian Disease ,firstly discovered in America whose pathogen is Aleutian Disease virus, was named by the mink with Aleutian coat color gene. It was demonstrated that all color phases of mink are susceptible to the disease although some differences in severity between the phases. Until now this disease has been spreaded throughout the world and became the the most important disease in mink.For this Disease there are several diagnostic assays, including the pathogens isolation, viral DNA amplification (PCR), iodine agglutination test (IAT), and ELISA spots note assay etc., but for many years considering the technical and experimental condition, CIE (CIEP) was used most widely in the world. Although this assay has a lot of advantages for example simplicity, less experimental condition demand and higher specificity, in recent years technology and testing standard improved this method more and more can not fulfil the requirements of mink breeding industry in China which has disadvantages of the big error of result determining, high possibility of spreading of virulent virus, long time consuming, so there is an urgent need for more safely, conveniently, fast assay.GICA is a new detection technology which based on cellulose nitrate membrane for solid-phase carrier, through the capillary action principle, in order to colloidal gold as a marker to show the results. This method has lots of advantages such as simplicity, low time consuming, clear result, easy-to-judgment, and small error, at the same time it does not need any equipment, low cost and doesn't need much experiences of technology for the operator, so this method is content for field and automated testing. Therefore, the technology has more and more fast development in vitro rapid test and veterinary.In this study against problems in Aleutian mink disease detection, apply with indirect colloidal gold immunochromatography assay (GICA), GICA for Aleutian disease of mink was established. Details of this study are following as:(1) In order to amplify specific fragments two pairs of specific primers against VP2 gene of the major epitopes are synthesized, the two fragments of amplification were cloned into the multiclone sites of prokaryotic expression vector pET-28a (+) and pMAL-p2X, then transform the recombinant plasmid into BL21 host bacteria, the two proteins from the samples collected by SDS-PAGE, Western-blot, can react with ADV specific antibody.(2) 25nm diameter colloidal gold particles are made by sodium citrate Reducting method and bath firing. Preparation of colloidal gold-labeled rabbit anti-mink IgG, to determine the best rabbit anti-mink IgG concentration is 12μg/mL, the optimal pH value is 9.1, used VP2A protein as a marker for detection of line, the optimal crossing concentration is 1mg/mL, used goat anti-rabbit IgG as a control line marker, the optimal crossing concentration is 0.8mg/mL.(3) According to the optimal labeling conditions test strips were assembled to determine sensitivity and specificity, and comparison with the immune electrical convection, showed that the sensitivity of GICA is 2 times higher than immune electrical convection, and no reaction with ADV-positive serum.
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