An optimized UPLC-MS/MS method for human plasma amyloid-β 42 and 40 measurement and application in Alzheimer's...

Critical events in Alzheimer's disease (AD) involve an imbalance between the production and clearance of amyloid-β (Aβ) peptides from the brain. The ratio of Aβ42 to Aβ40 in plasma was useful for evaluating AD, but quantification is limited by factors including preanalytical analyte loss and insufficient sensitivity. The availability of a targeted UPLC-MS/MS method with adequate analytical sensitivity and accurate values traceable to the SI units is essential for implementing a strategy for assay standardization. A targeted UPLC-MS/MS method for plasma Aβ42 and Aβ40 quantification was developed based on selected characteristic peptides spiked by ¹⁵N-labeled Aβ. The calibrator was assigned using an amino acid analysis reference method trace to SI units. UPLC-MS/MS conditions and sample preparation procedures were assessed. 59 plasma samples comparison were used to evaluate immunoassays. Additionally, two clinical cohorts were selected for diagnostic performance evaluation. The LOQ of Aβ42 and Aβ40 is 10 pg mL^-1 and 20 pg mL^-1, respectively. The linear range was 10-500 pg mL^-1 for Aβ42 and 20-1000 pg mL^-1 for Aβ40, recoveries between 95.3 % and 108.2 % for Aβ42, 93.2 % and 104.1 % for Aβ40, imprecisions were <7 %. The accuracy of method was validated by analysis of a certified reference material. Clinical cohorts for diagnostic performance evaluation shown that the area under the curve (AUC) for plasma Aβ42 and Aβ42/Aβ40 to differentiate between AD and CN were 0.767 and 0.799, respectively. A robust UPLC-MS/MS method was developed and demonstrated that suitable for a wide range of plasma Aβ42 and Aβ40. Applied to the investigation of clinically discrepant results, this method can act as an arbiter of the concentration of plasma Aβ42 and Aβ40 present.