This nuclear receptor assay system utilizes proprietary non-human mammalian cells engineered to provide constitutive, high-level expression of full-length, unmodified human androgen receptor (NR3C4), a ligand-dependent transcription factor commonly referred to as AR. The reporter cells include the luciferase reporter gene functionally linked to an AR-responsive promoter. Thus, quantifying changes in luciferase expression in the treated reporter cells provides a sensitive surrogate measure of the changes in AR activity. Luciferase gene expression occurs after ligand-bound AR undergoes nuclear translocation, DNA binding, recruitment and assembly of the co-activators and accessory factors required to form a functional transcription complex, culminating in expression of the target gene. Unlike some other cell-based assay strategies, the readout from The reporter cells demands the same orchestration of all intracellular molecular interactions and events that can be expected to occur in vivo. AR reporter cells are prepared using the proprietary CryoMite process. This cryo-preservation method yields high cell viability post-thaw, and provides the convenience of immediately dispensing healthy, division-competent reporter cells into assay plates. There is no need for intermediate spin-and-wash steps, viability determinations, or cell titer adjustments. The principal application of this assay product is in the screening of test samples to quantify functional activities, either agonist or antagonist, that they may exert against the human androgen receptor. It is an all-inclusive assay system that includes, in addition to AR reporter cells, two optimized media for use during cell culture and (optionally) in diluting the test samples, a reference agonist, luciferase detection reagent, a cell culture-ready assay plate, and a detailed protocol.
▪ AR Reporter Cells: 3 x 0.60 mL; -80°C▪ Cell Recovery Medium (CRM): 1 x 10.5 mL; -20°C▪ Compound Screening Medium (CSM): 1 x 35 mL; -20°C▪ 6α-Fl Testosterone, 10 µM (in DMSO)(reference agonist for AR): 1 x 30 µL; -20°C▪ Detection Substrate: 3 x 2.0 mL; -80°C▪ Detection Buffer: 3 x 2.0 mL; -20°C▪ Plate frame: 1; ambient▪ Snap-in, 8-well strips: 12; ambient