Ultrasensitive Assays Detect Different Conformations of Plasma β Amyloids

Abstract

With the developments of ultrasensitive technologies such as immunomagnetic reduction (IMR) assay, single molecule array (SIMOA) assay, electrochemiluminescence immunoassay (ECLIA), the assay of blood-based amyloid 1-42 (Aβ_1-42) becomes possible. However, the changes in measured plasma Aβ_1-42 concentrations in Alzheimer's disease (AD) compared to cognitively unimpaired subjects (CU) are inconsistent. A possible reason for the inconsistency regarding various conformations of Aβ_1-42 in plasma is explored in this study. Three samples with equal amounts of Aβ_1-42 but different proportions of monomers and oligomers of Aβ_1-42 were prepared. The Aβ_1-42 composition of monomers and oligomers in samples was analyzed with Western blot. Identically diluted versions of these three samples were assayed with IMR and SIMOA for Aβ_1-42 concentrations. The three diluted samples showed similar levels of Aβ_1-42 assayed with IMR, whereas much lower levels for samples with more oligomers assayed with SIOMA. The results imply that IMR detects both monomers and oligomers of Aβ_1-42. The measured levels of Aβ_1-42 are independent of the proportions of monomer or oligomer Aβ_1-42 but depend on the total amounts of Aβ_1-42. In the case of SIMOA, monomers of Aβ_1-42 are the primary target measured. By comparing Aβ_1-42 concentrations of the plasma using IMR and SIMOA, the significant difference in plasma Aβ_1-42 levels using IMR in AD compared to CU is mainly due to the formations of oligomeric Aβ_1-42. Therefore, if the target molecules are monomers of Aβ_1-42, SIMOA is the method of choice. Still, if the target molecules should include monomers, small and large oligomers, IMR would be an optimal consideration. In the future, the clinical implications of the proportion of oligomeric Aβ_1-42 need to be elucidated.

Figure 1

Detection of Aβ_1–42 monomer-rich and synthetic Aβ_1–42 oligomer-rich samples were detected by Western blotting. Lane 1: Aβ_1–42 monomer-rich sample (Sample 1); Lanes 2 and 3: Aβ_1–42 oligomer-rich samples (Samples 2, 3).

Figure 2

Measured Aβ_1–42 concentrations in d-Samples 1, 2, and 3 using IMR and SIMOA assays.

Figure 3

No significant correlation of plasma Aβ_1–42 concentrations between IMR and SIMOA measurements.

Figure 4

Schematic diagram shows how magnetic nanoparticles bind to monomers and oligomers in an immunomagnetic reduction assay. The magnetic particle binds to a monomer is smaller in size and lower in mass, thus having a higher spin velocity during oscillation. The magnetic particle binds to an oligomer is larger in size and higher in mass, consequently having a lower spin velocity during oscillation. The IMR analyzer applies an alternative current magnetic field to induce oscillation of all magnetic particles in the solution.