Fig. 1: Characterization of new pS129 and total α-synuclein SureFire Ultra assays using purified protein standards.
From: Novel tools to quantify total, phospho-Ser129 and aggregated alpha-synuclein in the mouse brain
AlphaLISA SureFire Ultra assays are designed using two antibodies against a target protein, which are differentially tagged to ensure their selective conjugation to one of two types of Alpha beads; donor beads or acceptor beads. The binding of both antibodies to an analyte brings donor and acceptor beads into very close proximity, which enables donor beads to activate acceptor beads following sample photoexcitation to produce the assay signal (a). Five new formulations of the pS129 (left) and total (right) α-synuclein assay were characterized using purified human (Hu) and mouse (Ms) pS129 α-synuclein standards (b–e), as well as human and mouse wild-type α-synuclein standards (WT; f–i). Data presented above represent only the most sensitive formulation of each assay. Associated data for all tested formulations are contained in Supplementary Tables 2, 3. Inter-assay variation (RSD) was calculated from three individual standard curves constructed and measured on separate days (c, e, i). The limit of detection (LoD) and lower limit of quantification (LLoQ) were defined as 3 and 6 standard deviations above the mean of the blank, respectively. Data in (b, d, f, h) represent mean ± standard deviation, while % variation data in (c, e, i) are only presented for standard concentrations detected above the LLoQ. For all standard curves; number of independent experiments (N) = 3, number of replicates in each experiment (n = 3). AU arbitrary units.
