Description
Overview
Endothelin receptors can be divided into ETA and ETB receptors, also called ETA1 and ETA2. Both receptors are involved in the treatment of hypertension.
This HEK293 stable cell line expresses the BETA2 receptor fused to a SNAP-Tag. Labeled with Terbium, the cells can be used in a Tag-lite receptor binding protocol.
Tag-lite assays offer many features and applications over traditional receptor/ligand binding assays:
Non-radioactive
Homogeneous and filtration-free
Ready-to-use kits and reagents for binding assays
Does not alter the receptor pharmacology
Peer-reviewed, validated technology
Thorough kinetic studies with true K
d
and K
i
values, association (K
on
) and dissociation (K
off
) rate constants
Screening and profiling of biologics and large molecules
How it works
Step 1 - Receptor labeling
SNAP-tag®, is a small fusion tag that covalently interacts with specific substrates. SNAP-tag allows specific and covalent labeling of any protein of interest. For more details, see the labeling procedure.
Step 2 - Understand the assay principle
Running a receptor binding assay using Tag-lite is as easy as it can get. Simply dispense 10 µL of labeled cells into each well, followed by 5 µL of labeled ligand and 5 µL of the compound you wish to test. Like all HTRF assays, Tag-lite assays do not require any washing steps. A diagram of the procedure to be followed is given on the right.
Step 3 - Saturation binding (KD)
A saturation binding assay measures total and non-specific binding for increasing concentrations of ligand under equilibrium conditions. To perform the assay, the fluorescent ligand is titrated into a solution containing a fixed amount of labeled cells and then incubated to equilibrium. The HTRF ratio obtained from this titration is the total binding.
Step 4 - Competitive binding (KI)
A competitive binding assay is performed to measure the dissociation constant, Ki. To perform the assay, the compound is titrated into a solution containing a fixed concentration of fluorescent ligand and a fixed amount of cells.
