应用化学发光法检测抗核抗体特定靶抗原/抗体的临床多中心研究

A multi-center clinical study for ANA specific autoantibodies detection by chemiluminescent immunoassay

目的 研究化学发光法（CLIA）检测抗核抗体（ANA）特定靶抗原/抗体的临床应用价值.方法 多中心研究.2016年4至7月对6家合作医院收集的811份标本,采用CLIA和免疫印迹法（LIA）开展抗核糖核蛋白（RNP）抗体、抗史密斯（Sm）抗体、抗干燥综合征抗原A（SSA/Ro60）抗体、抗干燥综合征抗原B（SSB/La）抗体、抗着丝点蛋白B（CENPB）抗体、抗双链DNA（dsDNA）抗体、抗核小体（Nuc）抗体和抗核糖体P蛋白（Rib-P）抗体的平行检测,比较两种方法抗体检出率、检测特异性以及检测符合率,对系统性红斑狼疮（SLE）高度特异性抗体（包括抗Sm、dsDNA、Nuc和Rib-P抗体）的差异样本采用酶联免疫吸附法（ELISA）进行复测,并结合SLE临床诊断信息进行McNemar检验分析.结果 CLIA与LIA在检测ANA特定靶抗原/抗体时,具有相当的抗体检出率和特异性.两种方法在检测抗RNP、SSA/Ro60、SSB/La和CENPB抗体时,表现出良好的一致性（Kappa〉0.75）,而在检测抗Sm、dsDNA、Nuc和Rib-P抗体时,一致性一般（0.4〈Kappa〈0.75）.抗Sm、dsDNA、Nuc和Rib-P这4个SLE特异性抗体的差异标本经ELISA复测,结果显示CLIA与ELISA在抗Sm抗体（χ^2=3.333,P=0.067）和抗Rib-P抗体（χ^2=0.888,P=0.345）检测结果差异无统计学意义.而LIA与ELISA在抗Sm抗体（χ^2=5.444,P=0.019）、抗dsDNA抗体（χ^2=5.812,P=0.015）、抗Nuc抗体（χ^2=12.071,P〈0.001）和抗Rib-P抗体（χ^2=25.861,P〈0.001）检测结果差异均具有统计学意义.结合差异标本诊断信息分析,结果显示CLIA抗dsDNA抗体（χ^2=1.132,P=0.249）和抗Nuc抗体（χ^2=0.571,P=0.449）的检测结果与SLE临床诊断差异无统计学意义,而LIA在抗Sm抗体（χ^2=21.125,P〈0.001）、抗dsDNA抗体（χ^2=59.507,P〈0.001）、抗Nuc抗体（χ^2=38.4,P〈0.001）和抗Rib-P抗体（χ^2=6.259,P=0.012）的检测结果与SLE临床诊断差异均存在统计学意义.结论 CLIA检测ANA特定靶抗原/抗体时与LIA具有相当的抗体检出率和特异性.两种方法在检测抗RNP、SSA/Ro60、SSB/La和CENPB抗体时具有良好一致性,但在检测抗Sm、dsDNA、Nuc和Rib-P抗体时一致性一般.对差异标本分析显示,CLIA结果与ELISA结果更有可比性,且CLIA结果与临床诊断信息更加符合.由于具备全自动、全定量、随机上样和灵活组合等显著特点,因此CLIA能够更好地满足ANA特定靶抗原/抗体定量检测的需求.

Objective To evaluate the clinical performance of chemiluminescent immunoassay （CLIA） on anti-nuclear antibody（ANA） specific autoantibodies testing.Methods A multi-center clinical study A total of 811 Sera samples were collected from 6 collaborating hospitals during the period of April to July 2016, and tested with CLIA and line immunoassay （LIA） in parallel for autoantibodies to ribonucleoprotein（RNP）, smith antigen（Sm）, SSA/Ro60,SSB/La, centromere protein B（CENPB）, double-stranded DNA（dsDNA）, nucleosome（Nuc）, and ribosome P protein（Rib-P）.The positive rate,specificity and qualitative coincidence rate for each antibody between CLIA and LIA methods were analyzed.All discrepant samples for systemic lupus erythematosus （SLE） highly specific autoantibodies （including anti-Sm, dsDNA, Nuc and Rib-P） were retested by enzyme linked immunosorbent assay （ELISA） and further analyzed with SLE disease cohort using McNemar test.Results The positive rate and specificity of CLIA and LIA for antibodies to ANA specific antigens were comparable.Excellent qualitative coincidence were found between CLIA and LIA for the detection of anti-RNP, SSA/Ro60, SSB/La and CENPB （Kappa〉0.75）, while the coincidence rate foranti-Sm, dsDNA, Nuc and Rib-P detection were moderate （0.4〈Kappa〈0.75）.The data from discrepant samples retested with ELISA showed there was no significant difference between CLIA and ELISA for the detection of anti-Sm （χ^2=3.333, P=0.067） and Rib-P （χ2=0.888, P=0.345）, but a significant difference were observed between LIA and ELISA test results for anti-Sm （χ^2=5.444, P=0.019）, anti-dsDNA （χ^2=5.812, P=0.015）, anti-Nuc （χ^2=12.071, P〈0.001） and anti-Rib-P （χ^2=25.861,P〈0.001）.When analyzing discrepant sample with SLE diagnosis, data from CLIA for anti-dsDNA （χ^2=1.132, P=0.249） and anti-Nuc （χ^2=0.571, P=0.449） showed no differencewith SLE diagnosis, while data from LIA for anti-Sm（χ^2=21.125,P〈0.001）, anti-dsDNA（χ^2=59.507,P〈0.001）, anti-Nuc（χ^2=38.4,P〈0.001） and anti-Rib-P （χ^2=6.259,P=0.012）showed significant difference with SLE diagnosis.Conclusions CLIA and LIA showed similar positive rate and specificity when testing antibody to ANA specific antigens.The coincidence rate between CLIA and LIA for the detection of anti-RNP, SSA/Ro60, SSB/La and CENPB wereexcellent and moderate for anti-Sm, dsDNA, Nuc and Rib-P.CLIA test results were more comparable with ELISA and had a better correlation with SLE disease diagnosis among all discrepant samples.With the additional benefits of full automation, quantitative output,random-access and high flexibility, CLIA is a preferable test platform for the detection of ANA specific autoantibodies.