Metrologically Traceable Quantification of 3 Apolipoprotein E Isoforms in Cerebrospinal Fluid

• Published in: Clinical chemistry (Baltimore, Md.) Md.), 2023-07, Vol.69 (7), p.734-745
• Main Authors: Huynh, Huu-Hien, Kuch, Kellie, Orquillas, Allen, Forrest, Katrina, Barahona-Carrillo, Lili, Keene, Dirk, Henderson, Victor W, Wagner, Anthony D, Poston, Kathleen L, Montine, Thomas J, Lin, Amy, Tian, Lu, MacCoss, Michael J, Emrick, Michelle A, Hoofnagle, Andrew N
• Format: Article
• Language: English
• Subjects: Alzheimer Disease - cerebrospinal fluid; Amyloid beta-Peptides - cerebrospinal fluid; Animals; Apolipoprotein E4 - cerebrospinal fluid; Apolipoprotein E4 - genetics; Apolipoproteins E - genetics; Cattle; Chromatography, Liquid; Humans; Protein Isoforms; Tandem Mass Spectrometry
• ISSN: 0009-9147; 1530-8561
• DOI: 10.1093/clinchem/hvad056

Abstract Background APOE genotype is associated with Alzheimer disease. Thus, the concentration of apolipoprotein E (apoE) isoforms in cerebrospinal fluid (CSF) could be altered in dementia. However, conflicting results have been obtained in different studies. Carefully validated and standardized assays could improve the interpretation of research findings, allow their replication in other laboratories, and generalize their application. Methods To evaluate this hypothesis, we aimed to develop, validate, and standardize a new measurement procedure using LC-MS/MS. Purified recombinant apoE protein standards (E2, E3, E4) were thoroughly characterized and used to assign the concentration of a matrix-matched calibration material that contained each apoE isoform, which ensured the metrological traceability of results. Results The assay of each isoform in human CSF was precise (≤11%CV) and of moderate throughput (approximately 80 samples per day). It demonstrated good linearity and parallelism for lumbar CSF, ventricular CSF, and bovine CSF. The use of an SI-traceable matrix-matched calibrator enabled precise and accurate measurements. There was no association observed between total apoE concentration and the number of Ɛ4 alleles in a cohort of 322 participants. However, the concentration of each isoform was significantly different in heterozygotes, with E4 > E3 > E2. Isoform concentrations were associated with cognitive and motor symptoms but contributed negligibly to a predictive model of cognitive impairment that included established CSF biomarkers. Conclusions Our method simultaneously measures each apoE isoform in human CSF with excellent precision and accuracy. A secondary matrix-matched material has been developed and is available to other laboratories to improve interlaboratory agreement.