Development Of Colloidal Gold Strip For Detection Of Antibody To Aleutian Mink Disease

Aleutian disease of mink(ADV)is caused by aleutian mink virus(ADV).The main characteristic of the chronic consumable and viral infection is increasing of plasma cells.It is mainly violate the immune system leading to the immune system to be disordered and caused extremely high death rate.The disease severely damage the mink farming industry vent this disease.The prevention and treatment of the disease has become a worldwide problem in the field of mink farming.There is no effective vaccine to prevent this disease.The prevention and treatment of the disease has become a worldwide problem in the field of mink farming.Therefore we mainly adopt the method of quarantine and eliminate positive minks to prevention and control the infection.In this study,the prokaryotic expression system and colloidal gold immunochromatography were used to investigate the rapid detection of paper strips by ADV.We used the biosoftware to analyze and intercepte the 705 bp fragment of the VP2 gene antigen epitopes of the ADV-DL125 strain isolated from the laboratory.Designed specific primers to amplify them by PCR.The prokaryotic expression vector pET-28a-VP2 a is constructed by the connect of PCR product with prokaryotic expression vector pET-28 a.pET-28a-VP2 a is transferred into the receptor BL21(DE3),and the specific protein bands can be obtained at about 28 KD after IPTG induced expression.The purified protein of VP2 a was obtained by Co-NTA affinity chromatography.The results of Western-blot showed that the purified protein of pET-28a-VP2 a could react well with the positive serum of ADV.We used the method of sodium citrate to prepare three different diameters of colloidal gold particles,and the colloidal gold particles at 30 nm were used to determine the labeling protein.According to the principle of gold immunochromatography,this study used VP2 a proteinas gold standard pad,the purified VP2 a protein as the NC film on the test line(T)of crossed antigen,and mink ADV positive serum IgG purified as a control line on the NC membrane(C)of antibody marking.By optimizing the reaction conditions,the best labeling concentration of gold labeled protein was10 mg / mL;the best marker pH was 8.69;T line antigen concentration was 0.7 mg/mL,C line antibody concentration as 0.9 mg/mL.The strip specificity,sensitivity,repeatability and meets the test results show that the development of the test strip react specifically with ADV positive serum of mink,mink distemper,and not with enteritis,PCV2 positive serum.counterimmunoelectrophoresis coincidence rate can reach 92.9%.