间接免疫荧光法在检测SARS冠状病毒特异性抗体中的应用

目的 利用间接免疫荧光技术(IFA)建立SARS冠状病毒(SARS—CoV)特异性抗体血清学诊断方法。方法 采用组织培养技术，用SARS病人的咽漱液样本，在Vero—E6细胞中培养，分离SARS冠状病毒；用已制备的SARS病毒抗原片与临床确诊为SARS的病人双相血清和正常人对照血清作用，再用荧光标记羊抗人IgM/IgG抗体与之结合，在荧光显微镜下观察荧光图像。结果 通过建立检测SARS冠状病毒特异性抗体的间接免疫荧光方法，在44份SARS临床病例标本中，检出IgG抗体阳性3l份，与临床诊断符合率为70．45％。31份IgG阳性者的急性期血清中IgM检出率为38．70％，IgM最早产生于发病第3天，发病7d以上的6例的IgM抗体均为阳性，滴度在发病12～15d达到高峰(1：80)；恢复期血清中IgG抗体最早产生于发病第8天，滴度在发病15～20d达到高峰(1：320～1：640)。97份正常人的血清标本中IgG抗体阳性率为3．09％，IgM抗体为阴性。结论 利用此方法，证实被SARS冠状病毒感染后，在机体中产生有特异性的IgM／IgG抗体；本方法检测结果与临床诊断符合率较高，可以用于早期临床诊断SARS病毒感染和辅助临床做出鉴别诊断。

IFA in testing specific antibody of SARS coronavirus

Objective To develop testing method for SARS coronavirus (SARS CoV) through IFA. Methods SARS CoVs were isolated from the Vero E6 cells inoculated with throat washings from 44 SARS patients. Antigen slides were prepared from infected cells in order to react with fluorescence labeled goat anti human IgM and IgG antibodies for further observation through fluorescence microscope. The procedure was controlled with 97 serum samples collected from healthy people. Results 31 of 44 patients were IgG positive through this approach, with coincident rate of 70.45% with clinical diagnosis. 38.70% of the IgG positives could be test positively of IgM in the acute phase, with the highest title of 1:80 presented during the 12-15 days of the course of the disease. The earliest time when the IgM could be detected was at 3 days after the onset and IgG in the convalescent sera could be firstly detected on 8 days of the course, with the highest title of 1:320-1:640 presented during 15-20 days of the course. However, the IgM was negative in all of the 97 control samples with only 3 IgG positive. Conclusion Specific IgM and IgG antibodies could be generated in the bodies after SARS CoV infection. The high coincident rate with clinic diagnosis showed the method can be applied in serologic test of SARS CoV infection to assist the clinic diagnosis.