Validation of beta-amyloid test as a reliable tool for quantifying beta-amyloid 1-40 and beta-amyloid 1-42 in blood

Abstract

Owing to the urgent requirement of easily accessible biomarkers for a wide-population screening of people at risk of suffering Alzheimer's disease (AD), there is increasing interest in blood AD biomarkers. The aim of this study was to validate two ELISA tests, ABtest40 and ABtest42, as reliable and sensitive tools for the quantification of Aβ1–40 and Aβ1–42 at levels as low as those present in blood. Inter and intra-assay precision was evaluated by repeated testing of 3 samples covering the whole range of quantification within 3 independent runs. Sensitivity of the assay was assessed by the estimation of the LLOQ (lower limit of quantification) and specificity was established by confirming absence of cross-reactivity with other Aβ species. Evaluation of relative error (RE) in quantification at LLOQ and ULOQ (upper limit of quantification) allowed the test accuracy assessment. Intra and inter-assay coefficients of variation were below 8% for both ABtest40 and ABtest42. The estimated LLOQ was 4.68pg/ml regarding ABtest40 and 11.95pg/ml for ABtest42. Accuracy of Aβ1–40 and Aβ1–42 quantification at LLOQ and ULOQ provided a relative error below 16% in all cases. Assays demonstrated very high affinity for their target analytes; cross reactivity was less than 3.35% for all the similar Aβ species tested at the ULOQ (N-terminal specificity assessed using Aβ2–40 and Aβ3–40 and C-terminal with Aβ1–38, Aβ1–43 and either Aβ1–40 or Aβ1–42). ABtest40 and ABtest42 proved to be accurate, specific and sensible enough to give a reliable quantification of low levels of Aβ1–40 and Aβ1–42.