Head-to-Head Comparison of 8 Plasma Amyloid-β 42/40 Assays in Alzheimer Disease

Key Points Question How well does plasma amyloid-β 42/40 (Aβ42/40), measured using 8 different assays, detect brain Aβ pathology in the early stages of Alzheimer disease? Findings In this study, including 408 participants from 2 independent cohorts (BioFINDER and Alzheimer Disease Neuroimaging Initiative), plasma Aβ42/40 quantified using certain mass spectrometry–based methods showed better discriminative accuracy than immunoassays when identifying individuals with abnormal intracerebral Aβ status according to cerebrospinal fluid Aβ42/40 levels and Aβ positron emission tomography. Meaning Certain mass spectrometry–based plasma tests might have sufficient performance to detect brain Aβ pathology in Alzheimer disease.

N-Aβ40 and 15 N-Aβ42 recombinant peptides were added to samples and Aβ peptides were immunoprecipitated with anti-amyloid-β antibodies 4G8 (epitope 17-27 in the amyloid-β sequence) and 6E10 (epitope 1-16, both antibodies from BioLegend). LC-MS/MS and analysis of mass spectrometry data were performed as previously described. 12 Analysis of Aβ40 and Aβ42 failed in 14 plasma samples, there was no second aliquot to repeat the analysis and, therefore, these samples were excluded from the present study. Plasma samples were analyzed at the Clinical Neurochemistry Laboratory, University of Gothenburg (Gothenburg, Sweden) in May-June 2021.

IA-Elc
CSF and plasma samples were analyzed using the Elecsys Aβ42 and Aβ40 immunoassays on a cobas e 601 analyzer at the Clinical Neurochemistry Laboratory, University of Gothenburg (Gothenburg, Sweden). CSF and plasma Aβ42 and Aβ40 assays were performed as previously described. 6 For plasma assays, different calibrator range and plasma controls were used. For plasma Aβ40 analysis, a biotinylated monoclonal Aβ40 specific antibody (23C2) and a monoclonal β-Amyloid-specific antibody (3D6) labeled with a ruthenium complex were used. The Elecsys β-Amyloid(1−42) CSF immunoassay is approved for use in countries accepting the CE mark; the Elecsys β-Amyloid(1−40) CSF immunoassay is a robust prototype assay (for research use only) which is not commercially available. Plasma samples were analyzed in November-December 2017.

IA-EI
CSF and plasma Aβ42 and Aβ40 were quantified using CSF and plasma kits according to the manufacturer's instructions. Samples were analyzed at EUROIMMUN AG (Luebeck, Germany) between May and November 2017.

IA-N4PE and IA-Quan
N4PE Simoa immunoassays are specific for the first amino acid of Aβ. These assays were developed by Amsterdam UMC and ADxNeurosciences and are now commercially available from Quanterix. 7,13 Plasma samples were analyzed using N4PE 4-plex kit according to the manufacturer's instructions. Plasma levels of Aβ42 and Aβ40 were also quantified using single-plex Simoa kits from Quanterix as previously described. 8 The Simoa Aβ40 and Aβ42 assays both utilize the same capture antibody targeting the N-terminus of βamyloid and different C-terminus detection antibodies specific to Aβ40 and Aβ42. The assays use β-amyloid (1-40) and (1-42) peptides as standards. The antibody pairs are the same in the IA-Quan Aβ42/Aβ40 singleplex kits and widely used Simoa Neuro 3-plex kit, but in the 3-plex kit the detection antibodies from the single-plex kits are used as capture antibodies and vice versa. IA-N4PE and IA-Quan analyses were performed at the Neurochemistry laboratory of the Amsterdam UMC location VUmc (Amsterdam, Netherlands) in May-June 2020 and Quanterix Corporation (Lexington, MA, USA) between December 2014 and January 2015, respectively.  14 PET/CT scanning of the brain was conducted at two sites using the same type of scanner (Gemini, Philips Healthcare, Best, the Netherlands). Sum images (from 90-110 min post injection) were analyzed using the software NeuroMarQ (GE Healthcare, Cleveland, OH, USA). [ 18 F]flutemetamol activity was quantified with a previously described fully automated PET-only method that uses an adaptive template for handling different uptake patterns in negative and positive [18F]flutemetamol images. 15 [ 18 F]flutemetamol images were spatially normalized to Montreal Neurological Institute template space using the adaptive template method. A volume of interest (VOI) template was applied for the following 9 bilateral regions: prefrontal, parietal, lateral temporal, medial temporal, sensorimotor, occipital, anterior cingulate, posterior cingulate/precuneus, and a global neocortical composite region composed by all these regions. 15 The standardized uptake value ratio (SUVR) was defined as the uptake in a VOI normalized for the cerebellar cortex uptake.

ADNI
ADNI plasma samples were analyzed between December 2020 and March 2021. Detailed information on sample handling procedures, assay protocols and performance are available from the ADNI database (for upto-date information, see www.adni-info.org and adni.loni.usc.edu). Aβ(1−40) Aβ(N-40) 3.1 11.0 0 * Analysis of Aβ40 and Aβ42 failed due to an over-pressure in the chomatographic system. ** ** One batch of assay was omitted due to unexpected system unstability Abbreviations: CV, coefficient of variability; IA, immunoassay; N/A, not available.      Aβ status was defined using previously described Aβ-PET cutoff (1.11). 17,18 . Biomarker concentrations are shown in pg/ml except Aβ42 IPMS-Shim and Aβ40 IPMS-Shim where the values are signal intensity normalized to the internal standard SIL-Aβ(1-38).