Concordance between APOE genotyping and proteotyping using the novel Lumipulse G Pan‐ApoE and Lumipulse G ApoE4 RUO assays

Abstract

Background: The APOE gene has three different alleles that encode for three different ApoE protein isoforms (APOE2, APOE3, and APOE4), resulting in 6 different phenotypes. Nucleic acid amplification test remains the gold standard for APOE genotyping, however quantification of the ApoE proteins using immunoassays could provide another piece of information, related to the expression levels of the proteins. In this regard, ApoE as a protein biomarker can be used in research to determine its value alone or as part of a panel of plasma biomarkers. Method: The LUMIPULSE G System is a chemiluminescent enzyme immunoassay platform enabling fully automated processing of samples using ready‐to‐use immunoreaction cartridges. Time to result in these single use test cartridges takes about 30 minutes. The analytical performance of the newly developed Lumipulse G Pan‐ApoE and Lumipulse G ApoE4 RUO assays was determined. Parameters analyzed were precision, sensitivity, linearity, prozone and potential impact of interfering substances. Proof of concept of these novel assays was verified with a set of 230 genotyped plasma samples. Result: The total within‐lab variability of both assays was ≤8% CV confirming high precision of the LUMIPULSE G system. Using low ApoE concentrated plasma samples, the observed LoD and LoQ (@20% CV) was 0.072 µg/mL (Pan‐ApoE) and 0.152 µg/mL (ApoE4). Linearity was shown across the assay range (0 – 100 µg/mL) for both assays. Neither prozone effect nor significant impact (recovery within 100±10%) of commonly tested endogenous substances (bilirubin, haemoglobin, low and high Protein content, and human anti‐mouse antibodies (HAMA)) was observed. Potential interference was observed for triglycerides and rheumatoid factor (RF) with observed recovery within 100±20%. ApoE proteotyping based on the novel assays resulted in a 99.8% concordance with APOE genotyping (1 discordant result). Moreover, applying the ratio ApoE4/Pan‐ApoE with the measurement values of respective assays, classified correctly between hetero‐ and homozygous E4 carrier status. Conclusion: The analytical performance studies demonstrate low variability and high sensitivity enabling measurement of pan‐ApoE and ApoE4 in plasma. Pan‐apoE and apoE4 quantification can offer relevant insights into the assessment of subjects at risk for AD or adverse events following anti‐amyloid treatments and should be studied further.