抗心磷脂抗体IgG时间分辨荧光免疫分析法的建立

Detection of anticardiolipin antibody IgG by time-resolved fluoroimmunoassay

目的建立高灵敏度、宽量程的定量检测患者血清中的抗心磷脂抗体(anticardiolipin antibody,ACA)IgG的时间分辨荧光免疫分析方法。方法采用ACA抗原(心磷脂+β2糖蛋白I)和鼠抗人IgG抗体分别作为固相抗原和铕标记抗体,如果样本中存在ACA抗体,则形成ACA抗原-ACA抗体-铕标记鼠抗人IgG抗体复合物,加入解离增强液解离铕离子,检测荧光强度,样本中ACA-IgG抗体含量与荧光强度成正比;对建立的时间分辨荧光免疫(time-resolved fluoroimmunoassay,TRFIA)法检测ACA抗体的线性范围、精密度、检测范围进行分析;对25例ACA-IgG抗体阳性血清标本分别利用TRFIA及ELISA法检测ACA-IgG抗体,分析其相关性;收集50例健康献血者,利用TR-FIA检测其血清中的ACA-IgG抗体,计算临床特异度。结果 利用SPSS 13.0统计软件进行统计学分析,TRFIA法检测高、中、低3种浓度混合血清的批内(n=20)精密度分别为2.34%、3.25%和3.87%,批间(n=8)精密度分别为2.89%、3.67%和4.50%;方法的灵敏度为0.1GPL U/ml;TRFIA法检测健康献血者,临床特异度为98%;TRFIA与ELISA 2种方法检测结果的一致性检验采用相关性分析,相关系数为0.987;TRFIA比ELISA的检测范围更宽,稳定性能好。结论首次建立稳定的高灵敏度和宽检测范围的TRFIA法检测人血清中的ACA-IgG抗体,对早期诊断自身免疫性疾病及监测疗效具有重要意义,该方法以其优势有望在各检验科室得以普遍应用。

Objective To establish a high sensitivity, wide range of quantitative detection of serum resistance to heart patients anticardiolipin antibody, prognosis （ACA） the IgG time-resolved fluorescence immunoassay analysis. method. Methods .Adopt ACA antigen （heart pbospbolipids ＋ beta 2 glycoprotein D and rat against human IgG respectively as solid-phase antigen antibody and eu mark antibody,If the sample ACA antibody, existing in ACA is formed ACA antibody-eu antigen-mark rat antiviral compounds,join antibody IgG disintegrate enhance liquid disintegrate euion,detection fluorescence intensity of ACA-IgG, sample antibody is proportional to the content and fluorescence intensity; To build a time-resolved fluorescence immunoassay （TRFIA）, resolved fluoroimmunoassay ACA antibody method to detect the linear range, precision,detection range analysis;25 cases of ACA-IgG antibody positive serum samples using TRFIA and ELISA respectively ACA-IgG antibody test,analyze the correlation;Collect 50 cases TRFIA detection by donors,health the ACA-IgG serum antibodies and clinical specific degrees calculation. Results use 13.0 statistical software SPSS statistical analyses,TRFIA method to detect the high,medium and low serum levels of three kinds of concentration of mixed （n = 20） within respectively 3.25% just 2.34% ,3. 25%,3.87%, Batch （n = 8） between respectively 3.67 % results, precision and 4.50% ;Methods sensitivity is 0.1 GPL U/ml;TRFIA method to detect health blood donors,clinical specific degrees for 98 percent respectively;With ELISA TRFIA testing results of the two methods by consistency inspection correlation analysis,correlation coefficient is 0. 987;The detection range than ELISA TRFIA more wide,stable performance. Conclusion First establish stable high sensitivity and wide range of testing method to detect the human serum TRFIA ACA- IgG antibody,Early diagnosis of autoimmune diseases and monitoring curative effect is of great significance his method t is expected to universal application.