背景:最近,用于诊断人类朊病毒疾病(HPD)的脑脊液(CSF)生物标志物的研究引起了极大的关注.在生物标志物研究中,重复性和准确性至关重要,特别是在总tau(T-tau)蛋白的测量中,这是一个重要的诊断标记。鉴于冠状病毒疾病大流行的全球影响,使用世界上一种全自动检测方法测量这种蛋白质的频率,化学发光酶免疫分析(CLEA),增加了。目前,神经系统疾病的诊断和监测主要依靠传统方法,但是它们的准确性和响应性有限。对tau测量中CLEA的准确性的了解有限。我们旨在使用CLEA测量T-tau蛋白并阐明其优点和局限性。
方法:我们随机选择60例快速进展性痴呆患者,采用ELISA和CLEA分析脑脊液标本。此外,我们使用Western印迹法检测14-3-3蛋白的存在,并使用实时震颤诱导转化(RT-QuIC)测定法分析同一组样品.此外,我们在60份样本中检测了ELISA和CLEA结果之间的相关系数.此外,使用CLEA,我们评估了昼夜重现性,储存稳定性,可稀释性,和冻融效应在三个选定的样品。
结果:在172名患者中,提取了172个样本,每个病人只提供一个样本,在RT-QuIC分析中,共有88名(35名男性和53名女性)的HPD检测呈阳性。相比之下,其余84例无HPD患者(50例男性和34例女性)的所有CSF样本在RT-QuIC分析中检测为阴性.ELISA和CLEA在测量T-tau蛋白水平方面均显示出完美的灵敏度和特异性(100%)。此外,ELISA和CLEA在测量灵敏度和检测极值的边际效应方面相似。CLEA分析显示某些T-tau蛋白水平超过2000pg/mL的样品不稳定。导致低重现性稀释分析。
结论:我们的发现表明CLEA在昼夜可重复性方面优于ELISA,储存稳定性,和冻融效应。然而,ELISA在稀释测定中表现出优异的性能。因此,在临床试验期间,必须开发用于CLEA测量的生物标志物样品稀释的创新方法.
BACKGROUND: Recently, the investigation of cerebrospinal fluid (CSF) biomarkers for diagnosing human prion diseases (HPD) has garnered significant attention. Reproducibility and accuracy are paramount in biomarker research, particularly in the measurement of total tau (T-tau) protein, which is a crucial diagnostic marker. Given the global impact of the coronavirus disease pandemic, the frequency of measuring this protein using one of the world\'s fully automated assays, chemiluminescent enzyme immunoassay (CLEA), has increased. At present, the diagnosis and monitoring of neurological diseases mainly rely on traditional methods, but their accuracy and responsiveness are limited. There is limited knowledge of the accuracy of CLEA in tau measurements. We aimed to measure T-tau protein using CLEA and to elucidate its merits and limitations.
METHODS: We randomly selected 60 patients with rapidly progressive dementia, using ELISA and CLEA analysis of cerebrospinal fluid specimens. Additionally, we used Western blotting to detect the presence of 14-3-3 protein and employed real-time quaking-induced conversion (RT-QuIC) assays to analyze the same set of samples. Furthermore, we examined the correlation coefficient between ELISA and CLEA results in a subset of 60 samples. Moreover, using CLEA, we evaluated the diurnal reproducibility, storage stability, dilutability, and freeze-thaw effects in three selected samples.
RESULTS: In 172 patients, 172 samples were extracted, with each patient providing only one sample, and a total of 88 (35 men and 53 women) tested positive for HPD in the RT-QuIC assay. In contrast, all CSF samples from the remaining 84 patients without HPD (50 men and 34 women) tested negative in the RT-QuIC assay. Both ELISA and CLEA showed perfect sensitivity and specificity (100%) in measuring T-tau protein levels. In addition, ELISA and CLEA are similar in terms of measurement sensitivity and marginal effect of detection extrema. CLEA analysis exhibited instability for certain samples with T-tau protein levels exceeding 2000 pg/mL, leading to low reproducibility during dilution analysis.
CONCLUSIONS: Our findings indicate that CLEA outperforms ELISA in terms of diurnal reproducibility, storage stability, and freeze-thaw effects. However, ELISA demonstrated superior performance in the dilution assay. Therefore, it is imperative to develop innovative approaches for the dilution of biomarker samples for CLEA measurements during clinical trials.