Human CAR-1 Reporter Assay Kit

SIZE	SKU	PRICE
1 x-96 well format assays	IB00911	—
3 x-32 assays in-96 well format	IB00911-32	—

SIZE	SKU
1 x-96 well format assays	IB00911
3 x-32 assays in-96 well format	IB00911-32

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Specifications
Data
Target Background
Documentation
Overview
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Product Description and Product Data

This is an all-inclusive cell-based luciferase reporter assay kit targeting the Human Constitutive Androstane Receptor-1 (CAR-1). INDIGO’s CAR-1 reporter assay utilizes proprietary mammalian cells that have been engineered to provide constitutive expression of the CAR-1. In addition to CAR-1 Reporter Cells, this kit provides two optimized media for use during cell culture and in diluting the user’s test samples, a reference agonist, Luciferase Detection Reagent, and a cell culture-ready assay plate. The principal application of this assay is in the screening of test samples to quantify any functional activity, either agonist or antagonist, that they may exert against human CAR-1. This kit provides researchers with clear, reproducible results, exceptional cell viability post-thaw, and consistent results lot to lot. Kits must be stored at -80C. Do not store in liquid nitrogen. Note: reporter cells cannot be refrozen or maintained in extended culture.

Features

Clear, Reproducible Results
All-Inclusive Assay Systems
Exceptional Cell Viability Post-Thaw
Consistent Results Lot to Lot

Product Specifications

Target Type		Nuclear Hormone Receptor
Species		Human
Receptor Form		Hybrid
Assay Mode		Agonist, Antagonist
Kit Components		CAR-1 Reporter Cells Cell Recovery Medium (CRM) Compound Screening Medium (CSM) PK11195, 15 mM (in DMSO) Detection Substrate Detection Buffer White, sterile, collagen-coated ready assay plate
Shelf Life		6 months
Orthologs Available		No
Shipping Requirements		Dry Ice
Storage temperature		-80C

Data

Dose-response analyses of the Human CAR1 Assay. Dose-response analyses of CAR1 Reporter Cells were performed using reference inverseagonist PK11195 (provided) according to the protocol provided in this Technical Manual. Reporter Cells were treated with seven concentrations (n ≥ 3). Average relative light units (RLU) +/- standard deviation (SD) were determined for each treatment concentration. Z’ values were calculated as described by Zhang, et al. (1999). Non-linear regression analyses and IC50 determination were performed using GraphPad Prism software. The high Z' values and fold-reductions in CAR1 activities confirm the robust performance of this CAR1 Reporter Assay System and demonstrate its suitability for use in HTS applications.

Target Background

The family of human Constitutive Androstane Receptors (CAR, NR1I3) regulate the expression of genes involved in xenobiotic metabolism and transport in the liver, including CYP2B and 3A4, UGT1 and MDR. Studies from mouse models show that CAR is also involved in bile acid, thyroid hormone and HDL homeostasis. The human CAR gene is subject to numerous alternative splicing events during pre-mRNA processing. The 348 amino acid isoform 1 of human CAR (CAR1) is encoded by 9 exons comprising the DNA binding domain (DBD), hinge region, and a ligand binding domain (LBD). The primary sequence of CAR2 differs from CAR1 in that it contains a four amino acid (VSPT) insert, whereas CAR3, which is the predominant isoform expressed in the liver, contains a distinct five amino acid (APYLT) insert.