Biomarkers

Abstract

Background: Amyloid β (Aβ) deposition in the brain is a pathological hallmark of Alzheimer's disease (AD). While immunoprecipitation-mass spectrometry (IP-MS) stands out as an accurate method for quantifying blood-based Aβ peptides, its major limitations such as prolonged sample preparation, extensive analysis time, large specimen volume, and high costs, present opportunities for improvement. Consequently, we aimed to develop a novel plasma IP-MS Aβ assay that employs simplified and significantly shorter analytical procedures, along with much-reduced sample volumes.

Method: We evaluated effects of reducing the sample volume, changing the buffers, implementing heavy labeled internal standards, and switching to a more cost-effective MALDI-Tof mass spectrometer versus the original Shimadzu IP-MS Aβ assay (Nakamura et al., 2018 Nature). This led to an improved assay, referred to as the Pittsburgh IP-MS Aβ assay. Analytical validation covered linearity, accuracy, precision, and recovery. In clinical validation, we compared the new and original Aβ assays for identifying Aβ-PET positivity and cognitive impairment in two independent cohorts.

Result: The Pittsburgh assay streamlined the immuno-enrichment step with a novel binding buffer, cutting background noise and reducing processing time by 50%. Additionally, it allowed for potential sample volumes as low as 50 µl and decreased antibody and paramagnetic bead usage by 75% each. The assay utilized the cost-friendly MALDI-Tof instrument from Bruker, a departure from the AXIMA Performance platform used in the original assay. The analytical validation showed consistent signal linearity, improved spike recovery with heavy labeled Aβ1-40 or Aβ1-42, but equivalent precision between the two IP-MS methods. In the AGUEDA cohort (n=148), the Pittsburgh assay demonstrated better AUCs than the older method for identifying Aβ-PET-positive individuals. This trend persisted in the University of Pittsburgh ADRC cohort (n=30) when comparing accuracy to identify patients with clinical assessed AD positive.

Conclusion: We have developed a more efficient and cost-effective IP-MS plasma Aβ assay, outperforming the original Shimadzu assay in clinical utility, while upholding consistently high analytical performance. The cost, time, and reagent savings along with the utilization of a more affordable and widely available instrument will empower more research laboratories to conduct IP-MS analysis of Aβ in blood more effectively.