摘要
以混合纤维素酯微孔滤膜为固相载体，用自制的禽流感全病毒抗原和酶标抗体，建立了禽流感抗体斑点 Ｅ Ｌ Ｉ Ｓ Ａ 检测法，其抗原最适包被量为 ０．０６μｇ／点；血清抗体最佳稀释度为 １１００；酶标抗体作 １２００ 稀释；出现明显清晰的斑点者判为禽流感抗体阳性。该方法对 Ｓ Ｐ Ｆ鸡血清及新城疫、传染性法氏囊病等其它 １１ 种鸡疫病阳性血清均为阴性，对不同亚型特异性的禽流感病毒（ Ａ Ｉ Ｖ）分型血清、琼扩（ Ａ Ｇ Ｐ）阳性血清及血凝抑制（ Ｈ Ｉ）阳性而 Ａ Ｇ Ｐ疑似的血清样品均呈阳性；对人工接种 Ａ Ｉ Ｖ 的 Ｓ Ｐ Ｆ鸡第 ３ 天即能检出抗体阳性，第 ５～１１７ 天可全部检出。与间接 Ｅ Ｌ Ｉ Ｓ Ａ 法比较，不仅其特异性、敏感性、重复性相一致，而且结果可用肉眼判定，更适合现地禽流感抗体监测及流行病学调查。
A Dot enzyme linked Immunosorbent assay(Dot ELISA) was established for detection of antibodies against influenza A virus(AIV)in chicken sera,employing the intact virus antigen and the enzyme labeled anti chicken Ig conjugate.The antigen was directly applied onto the mixed cellulose millipore filter with 0.06μg each dot for 15h at 4℃.The working titer of conjugate was 1200 dilution and that of serum samples were 1100 dilution.The samples with clear and obvious dots were judged as positive reaction for AIV antibody.The sera from specific pathogen free(SPF)and that from the chickens with 11 infectious diseases other than AI were negative in the Dot ELISA.However,the Dot ELISA could detect sera against all AIV subtypes which were positive for the antibody by agar gel precipitation(AGP)or by hemagglutinatin inhibition(HI)test.The sera from chickens infected with AIV strain A/Turkey/England/69(H3N2)were tested by Dot ELISA,AGP and HI tests.The anti AIV antibody could be detected 3 days post inoculation by ELISA,which was 2 and 7 days earlier than AGP and HI methods respectively.Comparing with indirect ELISA(iELISA),the Dot ELISA appeared correspondence in its specificity,sensitivity,and repeatability test whereas it's result was easily interpreted without an ELISA reader.The results above demonstrated that Dot ELISA was relatively practical an effective for antibody surveillance of avian influenza in chickens and could be widely adopted.
出处
《中国预防兽医学报》
CAS
CSCD
1999年第5期321-324,共4页
Chinese Journal of Preventive Veterinary Medicine
