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Analytical improvements in the detection of amyloid-β peptides in cerebrospinal fluid by mass spectrometry

University of British Columbia

Current diagnosis of Alzheimer’s disease (AD) is based on physical and neurological exams, mental state exams, and brain scans. Cerebrospinal fluid (CSF) biomarkers such as amyloid-β (Aβ) peptides have been incorporated into the research diagnostic criteria for AD and clinical care. Aβ peptides are produced from the amyloid precursor protein (APP) by proteolytic cleavage. Aβ40 and Aβ42, along with the protein tau, are reliable biochemical indicators of AD. Additionally autosomal dominant AD is caused from highly penetrant autosomal dominantly inherited variants in the APP, presenilin 1 and 2 genes. Biomarker detection and quantification is commonly performed by immunoassays, which provide an indirect measurement of the analyte of interest. In the case of Aβ immunoassays, high between-lab variability and lack of harmonization between platforms has been a concern. Moreover, immunoassays are known, in general, to be prone to analytical interferences – both positive and negative – from endogenous and exogenous molecules. To overcome issues associated with antibody-based detection methods and the desire to implement AD biomarker testing in routine care, a liquid chromatography tandem mass spectrometry (LC-MS/MS) assay was previously developed by the laboratory for detecting evidence of AD pathology by quantifying wild-type Aβ40 and Aβ42 in CSF. The aims of this thesis with respect to the LC-MS/MS method were to (1) evaluate the surrogate matrix composition, investigate potential interferences, and identify criteria for sample acceptance/rejection in a clinical environment, and (2) to adapt the method from detecting wild-type Aβ peptides to also detecting Aβ variants. The surrogate matrix was established with protein content compatible with that observed in human CSF. The interference studies indicated all endogenous and exogenous factors tested met the pre-specified bias acceptance criteria for Aβ40 and Aβ42. Thus Aβ peptides can be accurately quantified in the presence of high but physiologically plausible total protein concentrations and anti-Aβ antibodies, enabling use in routine clinical care and clinical trials. Additionally, testing hemolysate contaminated samples indicated no interference from hemolysate in the range commonly seen by the clinical laboratory. The LC-MS/MS assay was adapted to quantify wild type-Aβ42, Aβ40 and to identify Aβ variants in CSF.

Text

2020-08-31

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/71384

Pathology and Laboratory Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/