【作者】 李兴玉；

【导师】 文心田；

【作者基本信息】 四川农业大学 ， 预防兽医学， 2002， 硕士

【摘要】 本研究提取猪巴氏杆菌菌株的荚膜和外膜蛋白抗原，首次建立了检测猪巴氏杆菌病血清抗体的Dot-PPA-ELISA方法。本研究确定了抗原的最佳包被浓度为7.76ug／ml，酶标SPA的最佳稀释倍数为1：10。经动物试验初步确定了该方法的阳性标准。试验证明所建立的Dot-PPA-ELISA具有较好的特异性，与猪瘟、仔猪副伤寒、猪丹毒、猪细小病毒病、猪伪狂犬、猪布氏杆菌病、猪衣原体病阳性血清无交叉反应。试验所建立的Dot-PPA-ELISA灵敏度高，与琼扩试验比较，敏感度提高约128倍。诊断膜片具有很好的稳定性，4℃保存5个月，室温保存3个月，其特异性，灵敏度不变。本试验建立的Dot-PPA-ELISA方法操作方便、快速，结果客观、易于判定，重复性好，所用的相关试剂可以标准化。适用于养猪场猪巴氏杆菌病的诊断、免疫后抗体监测以及流行病学调查。更多还原

【Abstract】 A Dot-PPA-ELISA method has been developed to detect the antibody in swine serum against pasteurellosis. The capsule polysaccharide and outer membrane protein of pasteurella multocida was prepared as the diaphragm antigen. Applying the x2 test,optimal concentration of the diaphragm antigen was determined,and the optimum dilution of enzyme HRP-SPA 1:10. The high specificity of the Dot-PPA-ELISA was confirmed by specific blocking test and also by cross-reaction test. The diaphragm did not react with the antibodies against Salmonellosis,Streptococcosis,Colibacillosis,Chlamydiosis,HCV,PPV,PRV,Brucellosis. Erysipelas,suis and Chlamydiosis in cross-reaction test.The diaphragm has good sensitivity and could detect some pasteurella-positive test serum which has been diluted to2-". In comprision with GDPT ,the sensetivity is improved by 128 times . When 10 positive serum was detected 3 times,and the result was reproducible. The diaphragm has good stability as well,stored at 4 for at least 5 months or at 10-25 for 3 months,the sensitivity and specificity of the diaphragm did not change.This study endicates that the Dot-PPA-ELISA was a convenient,rapid,sensitive and specific method for the detection of antibody. Apart from these,all of the materials used in Dot-PPA-ELISA can be standardized. So it can be used for the surveillance of pasteurella multocida antibody in herd ,for the epidemiologic investigation of swine pasteurellosis ,and it provides a reference for a ratinal immunity schedule.更多还原