Korecka Magdalena, Waligorska Teresa, Figurski Michal, Toledo Jon B, Arnold Steven E, Grossman Murray, Trojanowski John Q, Shaw Leslie M
Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA Institute on Aging and Center for Neurodegenerative Disease Research, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
J Alzheimers Dis. 2014;41(2):441-51. doi: 10.3233/JAD-132489.
The primary aims of this work were to: 1) establish a calibrator surrogate matrix for quantification of amyloid-β (Aβ)42 in human cerebrospinal fluid (CSF) and preparation of quality control samples for LC-MS-MS methodology, 2) validate analytical performance of the assay, and 3) evaluate its diagnostic utility and compare it with the AlzBio3 immunoassay. The analytical methodology was based on a 2D-UPLC-MS-MS platform. Sample pretreatment used 5 M guanidine hydrochloride and extraction on μElution SPE columns as previously described. A column cleaning procedure involved gradual removal of aqueous solvents by acetonitrile assured consistent long-term chromatography performance. Receiver-operator characteristic (ROC) curve and correlation analyses evaluated the diagnostic utility of UPLC-MS-MS compared to AlzBio3 immunoassay for detection of Alzheimer's disease (AD). The surrogate matrix, artificial CSF containing 4 mg/mL of BSA, provides linear and reproducible calibration comparable to human pooled CSF as calibration matrix. Appropriate cleaning of the trapping and analytical columns provided every-day, trouble-free runs. Analyses of CSF Aβ42 showed that UPLC-MS-MS distinguished neuropathologically-diagnosed AD subjects from healthy controls with at least equivalent diagnostic utility to AlzBio3. Comparison of ROC curves for these two assays showed no statistically significant difference (p = 0.2229). Linear regression analysis of Aβ42 concentrations measured by this mass spectrometry-based method compared to the AlzBio3 immunoassay showed significantly higher but highly correlated results. In conclusion, the newly established surrogate matrix for 2D-UPLC-MS-MS measurement of Aβ42 provides selective, reproducible, and accurate results. The documented analytical performance and diagnostic performance for AD versus controls supports consideration as a candidate reference method.
1）建立用于定量人脑脊液（CSF）中淀粉样β蛋白（Aβ）42的校准替代基质，并制备用于液相色谱-串联质谱（LC-MS-MS）方法的质量控制样品；2）验证该检测方法的分析性能；3）评估其诊断效用，并将其与AlzBio3免疫分析方法进行比较。分析方法基于二维超高效液相色谱-串联质谱平台。样品预处理使用5M盐酸胍，并如前所述在μElution固相萃取柱上进行萃取。柱清洗程序包括通过乙腈逐步去除水性溶剂，以确保长期一致的色谱性能。接受者操作特征（ROC）曲线和相关性分析评估了与AlzBio3免疫分析相比，UPLC-MS-MS在检测阿尔茨海默病（AD）方面的诊断效用。替代基质，即含有4mg/mL牛血清白蛋白（BSA）的人工脑脊液，提供了与作为校准基质的人混合脑脊液相当的线性和可重复校准。对捕集柱和分析柱进行适当清洗可实现每日无故障运行。脑脊液Aβ42分析表明，UPLC-MS-MS能够将经神经病理学诊断的AD患者与健康对照区分开来，其诊断效用至少与AlzBio3相当。这两种检测方法的ROC曲线比较显示无统计学显著差异（p = 0.2229）。将基于这种质谱法测量的Aβ42浓度与AlzBio3免疫分析进行线性回归分析，结果显示前者显著更高但高度相关。总之，新建立的用于二维超高效液相色谱-串联质谱测量Aβ42的替代基质提供了选择性、可重复且准确的结果。所记录的针对AD与对照的分析性能和诊断性能支持将其视为候选参考方法进行考虑。
