Figure 1 | Scientific Reports

Figure 1

From: High-throughput screening against protein:protein interaction interfaces reveals anti-cancer therapeutics as potent modulators of the voltage-gated Na+ channel complex

Figure 1

Overview of the cell-based LCA for HTS against the FGF14:Nav1.6 C-tail complex. (a) Theory of LCA in live cells. Assembly of the CLuc-FGF14:CD4-Nav1.6-NLuc complex results in reconstitution of the luciferase enzymatic activity, which produces light in the presence of its substrate D-luciferin. (b) Linearized constructs encoding CLuc-FGF14-1b and CD4-Nav1.6-NLuc under the control of Neomycin and Puromycin, respectively, were sequentially transfected into HEK293 cells to create the double stable cell line. (c) Workflow for HTS using a cell-based assay. The work presented here describes assay development, screening and counter-screening of a test library of kinase inhibitors, and preliminary dose response hit validation. Z′ was used to measure the assay’s ability to detect hits, whereas Z-scores were calculated for experimental compounds based on per plate controls. CTB, CellTiter Blue (cell viability assay).

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