Developing novel supersensitive amyloid‐β 43 and pyroglutamate‐modified amyloid‐β 3‐40 assays for Alzheimer’s disease

Abstract Background In light of the poor fold change in the plasma amyloid‐β (Aβ) 42/amyloid‐β 40 ratio (Aβ 42/40 ) between Alzheimer’s disease (AD) and control (CTRL) cases, novel Aβ biomarkers are needed which will require harnessing the increased sensitivity offered by a novel modification to the Quanterix Corporation Single molecule array (Simoa) technology. Method Simoa assays for the detection of Aβ 43 and pyroglutamate‐modified Aβ 3‐40 (Aβ pE3‐40 ) have been developed on the standard HD‐X instrument. Aβ 43 and Aβ pE3‐40 were measured in 8 CSF samples (1 AD, 4 mixed phenotype [MP] and 3 CTRL) on the HD‐X instrument, each of which were measured at four dilution levels – neat, 2x, 4x and 8x dilution. Result Out of a possible 32 measurements per analyte (four dilution levels per sample), Aβ 43 was quantifiable in 68.8% of measurements, 72.7% of which fell below the concentration of calibrator B. In three samples, Aβ 43 was measurable at all four dilution levels, and showed a strong but statistically non‐significant dilution linearity. In contrast, Aβ pE3‐40 was quantifiable in 93.8% samples, 96.6% of which fell below the concentrations of calibrator B. In six samples, Aβ pE3‐40 was measurable at all four dilution levels. One sample showed a strong but statistically non‐significant dilution linearity, with the remaining five samples showing a poor dilution linearity, suggesting the limits of HD‐X sensitivity had been reached. Conclusion The increased sensitivity provided by the modification to the Simoa SRx is needed to accurately quantify Aβ 43 and Aβ pE3‐40 in CSF and subsequently plasma. Work is ongoing to transfer these novel assays from the HDx onto the modified SRx instrument.