MALDI‐TOF IP‐MS quantification of plasma amyloid peptides in Alzheimer’s disease

Abstract

AbstractBackgroundThe use of biomarkers which reflect Alzheimer’s disease (AD) improved the stratification of patients and the ability to propose personalized care. Their detection using either cerebrospinal fluid analysis (CSF), which is invasive, or using PET imaging, which is costly, is not adapted to large‐scale use. In 2018, Nakamura et al. using MALDI‐TOF IP‐MS generated a new composite biomarker with the amyloid‐ß precursor protein (APP)669–711/Aß42 and Aß40/Aß42 ratios with an AUC above 0.94 to identify subjects with PETAß+. Our primary objective here was to validate the use of this approach to identify AD and predicts its apparence in MCI and pre‐symptomatic patients.MethodThe blood‐based Aß markers were measured with an innovative MALDI‐TOF IP‐MS approach adapted from Nakamura et al., 2018. Briefly, 250 μl of plasma was diluted and spiked with 10 pM of stable‐isotope‐labelled (SIL) Aβ1–38 peptide. Aβ‐related peptides were immunoprecipitated, washed, and eluted with with 70% acetonitrile / 5 mM HCl to a 900‐μm μFocus MALDI plateTM (Hudson Surface Technology) which was pre‐spotted with a mix of α‐cyano‐4‐hydroxycinnamic acid (CHCA) and methanediphosphonic acid (MDPNA). Mass spectra were acquired using a MALDI‐linear TOF mass spectrometer (AXIMA Assurance, Shimadzu/KRATOS) equipped with a 337‐nm nitrogen laser in the positive ion mode. References AD cohorts with CSF biomarkers and longitudinal follow‐up were used.ResultsQuality controls (QC) plasma pools with Low (LQC), Medium (MQC), and High (HQC) (APP)669–711, Aß42 and Aß40 levels were generated to perform the analytical validation. Intra‐day variability were for LQC, MQC and HQC ≈19% for raw MS intensities and ≈6% after internal normalization. Inter‐day variability were ≈16% after normalization. Dilution integrity indicated linear results between 100% and 50% of the initial plasma volume. Three cycles of freeze/thaw test showed a good stability for the 3 target peptides. A cohort of >200 subject’s plasma samples from the memory consultation of the CMRR of Montpellier with Alzheimer’s disease, Mild Cognitive Impairment and other degenerative diseases are being analyzed to demonstrate the clinical interest of this measurement.ConclusionMALDI‐TOF IP‐MS quantification of plasma amyloid peptides reached satisfactory analytical performance for clinical use. Its relevance for Alzheimer disease will be illustrated.