Rat Androgen Receptor (rAR; nr3c4)
|Product Family||Product Number||Product Description||Technical Manual|
|R03001-32||Rat AR Reporter Assay System, 3 x 32 assays in 96-well format||Technical Manual|
|R03001||Rat AR Reporter Assay System, 1 x 96-well format assays||Technical Manual|
Rat Androgen Receptor Assay Kit
This Rat Androgen Receptor (rAR) assay kit is an all-inclusive firefly luciferase reporter assay system that includes in addition to rAR 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.
Rat AR Reporter Cells are prepared using INDIGO’s 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.
INDIGO’s assay kits feature a luciferase detection reagent specially formulated to provide stable light emission between 5 and 90+ minutes after initiating the luciferase reaction. Incorporating a 5-minute reaction-rest period ensures that light emission profiles attain maximal stability, thereby allowing assay plates to be processed in batch. By doing so, the signal output from all sample wells, from one plate to the next, may be directly compared within an experimental set.
Kits are offered in different assay formats to accommodate researchers' needs: 3x 32 and 1x 96 assay formats for screening small numbers of test compounds, as well as custom bulk reagents for HTS applications.
Bulk assay reagents can be custom manufactured to accommodate any scale of HTS. Please inquire.
Rat Androgen Receptor Assay Services
The primary application of INDIGO's cell-based nuclear receptor assays are to quantitatively assess the bioactivity of a test compound as an agonist (activator) or antagonist (inhibition of an agonist response) of a given receptor. Service assays include a positive control reference compound and 'vehicle' control for every experiment. A formal study report and all data files are provided to the client upon completion of the study. To receive a quote for your proposed study, complete & submit the online "Request a Quote" form or contact an INDIGO Customer Service Representative to discuss your desired study parameters.
Rat Androgen Receptor Assay Background
The androgen receptor is activated by binding of either of the androgenic hormones testosterone or dihydrotestosterone. The androgen receptor is most closely related to the progesterone receptor, and progestins in higher dosages can block the androgen receptor. The main function of the androgen receptor is as a DNA binding transcription factor which regulates gene expression; however, the androgen receptor has other functions as well.
Androgen regulated genes are critical for the development and maintenance of the male sexual phenotype. In some cell types testosterone interacts directly with androgen receptors while in others testosterone is converted by 5-alpha-reductase to dihydrotestosterone, an even more potent agonist for androgen receptor activation. Testosterone appears to be the primary androgen receptor activating hormone in the Wolffian duct while dihydrotestosterone is the main androgenic hormone in the urogenital sinus, urogenital tubercle, and hair follicles. Hence testosterone is primarily responsible for the development of male primary sexual characteristics while dihydrotestosterone is responsible for secondary male characteristics.
This nuclear receptor assay system utilizes proprietary non-human mammalian cells engineered to provide constitutive, high-level expression of full length, unmodified Rat Androgen Receptor (nr3c4; AR), a ligand-dependent transcription factor commonly referred to as rAR.
INDIGO’s Reporter Cells include the luciferase reporter gene functionally linked to an AR-responsive promoter. Therefore, quantifying changes in luciferase expression in the treated reporter cells provides a sensitive surrogate measure of the changes in rAR activity. Luciferase gene expression occurs after ligand-bound rAR 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 INDIGO’s reporter cells demands the same orchestration of all intracellular molecular interactions and events that can be expected to occur in vivo.
The principle 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 rat androgen receptor.
For more information on AR, visit the Nuclear Receptor Resource.
Rat Androgen Receptor Assay Data
Rat AR Agonist dose-response analyses. Rat AR agonist dose-response assays were performed using the reference agonists 5αDihydro 11-keto Testosterone (5αDH-11kT; provided), Dihydro Testosterone (DHT; Steraloids Inc.), 6α-Fl Testosterone (6αFlT; Enzo Life Sci), Cl-4As-1, 11-keto Testosterone (11-kT; Sigma-Aldrich) and Medroxy-Progesterone 17-Acetate (MPA; Steraloids, Inc.). Luminescence was quantified and values of average relative light units (RLU), corresponding standard deviation (SD), Fold-Activation and Z’ were determined. Non-linear regression analyses of Fold-Activation vs. Log10 transformed concentration were plotted and EC50 values determined using GraphPad Prism software.