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粪肠球菌抗体间接ELISA检测方法的建立及应用
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周霞，王晓兰，剡根强，马勋
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（石河子大学 动物科技学院,新疆 石河子832003）
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摘要：
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利用超声波裂解法制备粪肠球菌的裂解物作为抗原包被酶标反应板，通过棋盘滴定方法筛选最佳反应条件，成功地建立了检测致羔羊脑炎粪肠球菌抗体的间接ELISA。在此间接ELISA中，抗原最佳包被浓度为37.57μg/mL，被检血清的最佳稀释度为1∶100。建立的间接ELISA的灵敏度是微量凝集反应的25～100倍。交叉试验、阻断试验、重复性试验表明，该方法具有重复性好、特异性强、操作简便快速等特点。确立的检测ELISA效价的回归方程lg\[1/ET\]=1.275+2.730 D405nm,可用于抗体定量测定。利用此方法检测了来自未免疫绵羊场的50份成年绵羊的血清和50份羔羊的血清；结果，2份成年绵羊的血清为阳性，其他样品均为阴性。 |
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关键词：
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间接ELISA；粪肠球菌；羔羊；脑炎 |
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中图分类号：S 855.11 文献标识码：
A 文章编号：1673-4696(2008)05-0417-05 |
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Development and application of indirect ELISA for detection of antibody
against Enterococcus faecalis |
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ZHOU Xia，WANG Xiao-lan，YAN Gen-qiang,MA Xun |
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(College of Animal Science and Technology，Shihezi University,Shihezi 832003，China) |
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Abstract: An indirect ELISA for detecting the antibodies against Enterococcus faecalis causing lamb encephalitis was developed successfully by phalanx titration with coating antigen from E.faecalis cracked by ultrasonic.The optimal working concentration of the antigen was 37.57μg/mL，and the optimal dilution of the serum to be detected was 1∶100 in the indirect ELISA.The indirect ELISA was 25-100 times sensitive than micro-agglutination test.The crossing-test，reproducibility test and blocking-test showed that the indirect ELISA was more specific，easier and quicker.The titers of the antibody could be determined quantitatively by the regression equation lg\[1/ET\]=1.275+2.730 D405nm.50 adult ovine sera and 50 lamb sera from the unimmunized farms were examined by the indirect ELISA,and 2 positive sera were detected in 50 adult sheep sera but no positive serum in 50 lamb sera. |
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Key words: indirect ELISA;Enterococcus faecalis；lamb;encephalitis |
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