A3R expressing cells labeled with Terbium
Tag-lite cells transiently expressing the A3 receptor with Terbium
The Tag-lite A3 Adenosine receptor plasmid is used to transiently or stably transfect cells in order to develop an A3 Adenosine receptor binding assay.
Over the past few years, SNAP-Tag technology combined with TR-FRET has paved the way for the development of many non-radioactive, no-wash, binding assays. The method is based on transfecting cells using plasmids encoding a SNAP-Tag and subsequently labeling them with Terbium. Cisbio offers a large collection of such plasmids. All GPCR genes are cloned in an expression vector directly downstream from a CMV promoter, and are provided ready for protein expression and labeling.
All information on this page pertains to the Tag-lite plasmid cloned with the Adenosine A3 receptor.
SNAP-tag® is a small fusion tag that covalently interacts with specific substrates. It enables the specific and covalent labeling of any protein of interest (refer to labeling procedure). Cells are provided unlabeled, and need to be labeled with Lumi4-Terbium prior to running a binding assay. Labeling reagents are available in 4 different sizes from the Cisbio catalog.
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.
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.
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