Alzheimers Disease (AD) biomarker measurement is key to aid in the diagnosis and prognosis of the disease. In the research setting, participant recruitment and retention and optimization of sample use, is one of the main challenges that observational studies face. Thus, obtaining accurate established biomarker measurements for stratification and maximizing use of the precious samples is key. Accurate technologies are currently available for established biomarkers, mainly immunoassays and immunoprecipitation liquid chromatography-mass spectrometry (IP-MS), and some of them are already being used in clinical settings. Although some immunoassays- and IP-MS based platforms provide multiplexing for several different coding proteins there is not a current platform that can measure all the stablished and emerging biomarkers in one run. The NUcleic acid Linked Immuno-Sandwich Assay (NULISA) is a mid-throughput platform with antibody-based measurements with a sequencing output that requires 15uL of sample volume to measure more than 100 analytes, including those typically assayed for AD. Here we benchmarked and compared the AD-relevant biomarkers including in the NULISA against validated assays, in both CSF and plasma. Overall, we have found that CSF measures of Abeta42/40, NfL, GFAP, and p-tau217 are highly correlated and have similar predictive performance when measured by immunoassay, mass-spectrometry or NULISA. In plasma, p-tau217 shows a performance similar to that reported with other technologies when predicting amyloidosis. Other established and exploratory biomarkers (total tau, p-tau181, NRGN, YKL40, sTREM2, VILIP1 among other) show a wide range of correlation values depending on the fluid and the platform. Our results indicate that the multiplexed immunoassay platform produces reliable results for established biomarkers in CSF that are useful in research settings, with the advantage of measuring additional novel biomarkers using minimal sample volume.

RB has received funding from NIH, Alzheimers Association, Biogen, AbbVie, Bristol Meyer Squibbs, Novartis, and EISAI. RB has equity and is on the scientific advisory board of C2N Diagnostics. SES has served on advisory boards and consulted on biomarker testing for Eisai, and she has received speaker fees from Eli Lilly. DMH has equity and is on the scientific advisory board of C2N Diagnostics. DMH is on the scientific advisory board of Denali, Genentech, and Cajal Neuroscience and consults for Asteroid. TLSB received research funding from Siemens and is an investigator on clinical trials and studies of AD with partial support from Eisai, Lilly, Roche, Janssen, and Biogen. TLSB is a consultant to Eisai. CC has received research support from: GSK and Eisai. CC is a member of the advisory board of Circular Genomics and owns stocks. CC is a member of the advisory board of ADmit. The other authors report no conflict of interest. The funders of the study had no role in the collection, analysis, or interpretation of data; in the writing of the report; or in the decision to submit the paper for publication.

This work was supported by grants from the National Institutes of Health (R01AG044546 (CC), P01AG003991(CC, JCM), RF1AG053303 (CC), RF1AG058501 (CC), U01AG058922 (CC), K99/R00-AG062723 (LI), RF1AG074007 (YJS), R01AG070941 (SES)), the Chan Zuckerberg Initiative (CZI), the Michael J. Fox Foundation (LI, CC), the Department of Defense (LI- W81XWH2010849), the Alzheimers Association Zenith Fellows Award (ZEN-22-848604, awarded to CC), the Alzheimer's Drug Discovery Foundation (LI), Bright Focus Foundation (LI), and an Anonymous foundation. The recruitment and clinical characterization of research participants at Washington University were supported by NIH P30AG066444 (JCM), P01AG03991(JCM), and P01AG026276(JCM). This work was supported by access to equipment made possible by the Hope Center for Neurological Disorders, the Neurogenomics and Informatics Center (NGI: https://neurogenomics.wustl.edu/) and the Departments of Neurology and Psychiatry at Washington University School of Medicine.

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The details of the IRB/oversight body that provided approval or exemption for the research described are given below:

All research was performed in accordance with the protocols approved by the Institutional Review Board (IRB) at Washington University in Saint Louis.

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I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).

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All data produced in the present study are available upon reasonable request to the authors