目的:血液中胶质纤维酸性蛋白(GFAP)水平升高已被确定为某些神经系统疾病的有价值的生物标志物,如阿尔茨海默病和多发性硬化症。然而,到目前为止,大多数血液GFAP定量都是使用相同的基于珠子的测定法进行的,迄今为止,缺乏常规的临床应用。
方法:在本研究中,我们验证了一种新的第二代(第二代)Ella测定法来定量血清GFAP.此外,我们在神经系统疾病的临床队列中,将其性能与基于珠子的单分子阵列(Simoa)和自制GFAP测定进行了比较,包括210名患者。
结果:验证实验产生10%的测定内变异,12%的中间试验,检测限为0.9pg/mL,定量下限为2.8pg/mL,暴露于五个冻融循环的血清样本变化小于20%,在4°C和室温下120h。使用所有测定的临床群组的测量揭示了不同诊断组中相同的GFAP分布模式。此外,我们观察到第二代Ella和Simoa之间存在很强的相关性(r=0.91(95%CI:0.88-0.93),p<0.0001)和自制免疫测定(r=0.77(95%CI:0.70-0.82),p<0.0001)。
结论:我们的结果证明了高可靠性,第2代Ella测定的精密度和可重复性。尽管观察到对Simoa的检测灵敏度更高,这种新的微流控检测方法由于其稳定性和易用性,可能有可能在日常临床检查中用于GFAP分析.
OBJECTIVE: Increased levels of glial fibrillary acidic protein (GFAP) in blood have been identified as a valuable biomarker for some neurological disorders, such as Alzheimer\'s disease and multiple sclerosis. However, most blood GFAP quantifications so far were performed using the same bead-based assay, and to date a routine clinical application is lacking.
METHODS: In this study, we validated a novel second-generation (2nd gen) Ella assay to quantify serum GFAP. Furthermore, we compared its performance with a bead-based single molecule array (Simoa) and a homemade GFAP assay in a clinical cohort of neurological diseases, including 210 patients.
RESULTS: Validation experiments resulted in an intra-assay variation of 10 %, an inter-assay of 12 %, a limit of detection of 0.9 pg/mL, a lower limit of quantification of 2.8 pg/mL, and less than 20 % variation in serum samples exposed to up to five freeze-thaw cycles, 120 h at 4 °C and room temperature. Measurement of the clinical cohort using all assays revealed the same pattern of GFAP distribution in the different diagnostic groups. Moreover, we observed a strong correlation between the 2nd gen Ella and Simoa (r=0.91 (95 % CI: 0.88-0.93), p<0.0001) and the homemade immunoassay (r=0.77 (95 % CI: 0.70-0.82), p<0.0001).
CONCLUSIONS: Our results demonstrate a high reliability, precision and reproducibility of the 2nd gen Ella assay. Although a higher assay sensitivity for Simoa was observed, the new microfluidic assay might have the potential to be used for GFAP analysis in daily clinical workups due to its robustness and ease of use.