大鼠肺炎支原体感染的ELISA实验研究

Study on the ELISA Diagnosis of Rats infected with M.Pneumonia

目的探讨支原体感染大鼠的血清抗体反应及其间接ELISA检查与诊断方法。方法将肺炎支原体疫苗分别以滴鼻、腹腔注射、尾静脉注射方式免疫大鼠,收集动物血清,以方阵滴定法和间接法确立ELISA检测实验的最适条件,用间接法检测不同方法与时间免疫动物的血清特异性IgG抗体。结果ELISA系统的最适工作条件包括抗原包被浓度10×10^(-6)g/ml、血清稀释度1:40、酶结合物1:5000。以该系统检测滴鼻免疫大鼠共21只,免疫6次2只大鼠血清抗体阳性(66．67%),免疫7次后阳性率达100%;腹腔免疫和静脉免疫大鼠各18只,免疫2次后2只大鼠可检出特异性抗体(66．67%),免疫5次后全部动物均为血清抗体阳性(100%)。结论肺炎支原体0．0016 g/只以滴鼻、腹腔注射、尾静脉注射的方法均能够使大鼠产生特异性抗体,以间接ELISA检测法检查腹腔免疫组和静脉免疫组大鼠血清免疫2次后即有部分大鼠血清抗体检测即呈阳性,5次后阳性率达100%,而滴鼻免疫组大鼠直至免疫6次后部分血清抗体检测才可呈阳性其阳性率与腹腔免疫和静脉免疫2次阳性率相当。间接ELISA法可有效检测支原体感染动物的血清特异性抗体,是一种具有较高特异性的快速简便方法。

Objective To study the antibody response in the sera of rats infected by M. pneumonia and to probe the method of ELISA diagnosis. Methods The rats were infected separately with M. pneumonia by intranasal, intraperitoneal or intravenous and then the sera were collected at various times. The specific antibodies in the samples were tested separately with the indirect ELISA. Results The most suitable one of reactive condition was determined that including the antigen for coating of 10 ×10^-6 g/ml, the sera of 1 ： 40 and the enzyme conjugate of 1 ：5 000. In 21 rats with intranasal immune, positive reaction was found in 2 animals after 6^th times of immune（66. 67%）, and positive reaction in all of the rats was showed after 7^th times of immune. In 18 rats with intraperitoneal immune, there were 2 animals showed positive reaction after 2^nd times of immune（66. 67%）and all of the rats were positive reaction after 5^th times of immune. In the group with intravenous immune, the results were same as that of the group with intraperitoneal immune. Conclusion The specific antibodies could be produced in sera of rats by intranasal, intraperitoneal injection or intravenous injection with M. pneumonia. The antibodies could be detected after 2 times of immune but higher positive rate would be got until 5 or 6 times. Indirect ELISA is a useful method with quick, sensitive, specific and easy to diagnosis of Mycoplasma infection of rat.