Development and technical validation of an immunoassay for the detection of app669–711 (Aβ−3–40) in biological samples

Klafki HW, Rieper P, Matzen A, Zampar S, Wirths O, Vogelgsang J, Osterloh D, Rohdenburg L, Oberstein T, Jahn O, Beyer I, Lachmann I, Knölker HJ, Wiltfang J (2020)

Publication Type: Journal article

Publication year: 2020

Journal

International Journal of Molecular Sciences MDPI

Book Volume: 21

Pages Range: 1-25

Article Number: 6564

Journal Issue: 18

DOI: 10.3390/ijms21186564

Abstract

The ratio of amyloid precursor protein (APP)669–711 (Aβ−3–40)/Aβ1–42 in blood plasma was reported to represent a novel Alzheimer’s disease biomarker. Here, we describe the characterization of two antibodies against the N-terminus of Aβ−3–x and the development and “fit-for-purpose” technical validation of a sandwich immunoassay for the measurement of Aβ−3–40. Antibody selectivity was assessed by capillary isoelectric focusing immunoassay, Western blot analysis, and immunohistochemistry. The analytical validation addressed assay range, repeatability, specificity, between-run variability, impact of pre-analytical sample handling procedures, assay interference, and analytical spike recoveries. Blood plasma was analyzed after Aβ immunoprecipitation by a two-step immunoassay procedure. Both monoclonal antibodies detected Aβ−3–40 with no appreciable cross reactivity with Aβ1–40 or N-terminally truncated Aβ variants. However, the amyloid precursor protein was also recognized. The immunoassay showed high selectivity for Aβ−3–40 with a quantitative assay range of 22 pg/mL–7.5 ng/mL. Acceptable intermediate imprecision of the complete two-step immunoassay was reached after normalization. In a small clinical sample, the measured Aβ42/Aβ−3–40 and Aβ42/Aβ40 ratios were lower in patients with dementia of the Alzheimer’s type than in other dementias. In summary, the methodological groundwork for further optimization and future studies addressing the Aβ42/Aβ−3–40 ratio as a novel biomarker candidate for Alzheimer’s disease has been set.

Authors with CRIS profile

Timo Oberstein Department of Psychiatry and Psychotherapy

Involved external institutions

Universitätsklinikum Göttingen DE Germany (DE) Tecan Group Ltd. CH Switzerland (CH) AJ Roboscreen GmbH DE Germany (DE) Max-Planck-Institut für Experimentelle Medizin DE Germany (DE) Technische Universität Dresden DE Germany (DE)

How to cite

APA:

Klafki, H.W., Rieper, P., Matzen, A., Zampar, S., Wirths, O., Vogelgsang, J.,... Wiltfang, J. (2020). Development and technical validation of an immunoassay for the detection of app669–711 (Aβ−3–40) in biological samples. International Journal of Molecular Sciences, 21(18), 1-25. https://doi.org/10.3390/ijms21186564

MLA:

Klafki, Hans W., et al. "Development and technical validation of an immunoassay for the detection of app669–711 (Aβ−3–40) in biological samples." International Journal of Molecular Sciences 21.18 (2020): 1-25.

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