| Product Description |
Recombinant CHO-K1 cells stably overexpress human nicotinic acid receptor (NIACR1), also known as hydroxycarboxylic acid receptor 2 (HCAR2), on the surface and contain high levels of G protein Gαi to couple with the receptor in downstream signaling pathways. |
| Applications |
Calcium mobilization assay and cAMP accumulation assay. |
| Expressed Gene |
NCBI reference sequence NM_177551; no expressed tags |
| Target Protein |
NP_808219 |
| Host Cell |
CHO-K1 |
| Size |
Two vials of frozen cells (>1×106 per vial in 1 mL) |
| Storage |
Store cells in liquid nitrogen immediately upon receipt. Thaw and recover cells within one year from the date received. |
| Mycoplasma Status |
Negative. The mycoplasma test was performed with MycoAlert™ PLUS Mycoplasma Detection Kit (Cat. No. LT07-318, Lonza). |
| Stability |
Stable through more than 16 passages with no significant changes in assay performance or expression profile. |
| Culture Properties |
Adherent |
| Freeze Medium |
45% Ham’s F-12K (Kaighn’s) (Cat. No. 21127, Life Technologies), 45% FBS (Cat. No. 10099-141, Gibco), 10% DMSO (Cat. No. D2650, Sigma) |
| Complete Growth Medium |
Ham’s F-12K (Kaighn’s), 10% FBS |
| Culture Medium |
Ham’s F-12K (Kaighn’s), 10% FBS, 200 μg/ml Zeocin (Cat. No. R250-01, Life Technologies), 100 μg/ml Hygromycin B (Cat. No. 10687010, Invitrogen) |
| Gene Synonyms |
HCAR2; HCA2; HM74a; HM74b; PUMAG; Puma-g; GPR109A |
| Background |
GPR109A is a high affinity receptor for nicotinic acid (niacin) and is a member of the nicotinic acid receptor family of G protein-coupled receptors (the other identified member being GPR109B). GPR109A is a Gi/o protein-coupled receptor with high affinity for nicotinic acid. In GPR109A knockout mice, the effects of niacin on both lipids and flushing are eliminated. Furthermore in arrestin beta 1 knockout mice, niacin's effect on flushing is greatly reduced while the lipid modifying effects are maintained. GPR109A is believed to be an important biomolecular target of niacin which is a widely prescribed drug for the treatment of dyslipidemia and to increase HDL cholesterol but whose therapeutic use is limited by flushing. |
Figure 1. Nicotinic acid-induced concentration-dependent stimulation of intracellular calcium mobilization in CHO-K1/NIACR1/Gα15 cells. The cells were loaded with Calcium-4 (Cat. No. R8142; Molecular Devices) prior to stimulation with NIACR1 agonist, nicotinic acid. The intracellular calcium change was measured by FLIPRTETRA. The relative fluorescent units (RFU) were recorded and normalized to plot against the log of the cumulative doses of nicotinic acid (mean ± SEM, n = 3). The EC50 of nicotinic acid on CHO-K1/NIACR1/Gα15 cells was 1.38 nM.
Notes:
EC50 value is calculated with four parameter logistic equation:
Y=Bottom + (Top-Bottom) / (1+10^ ((LogEC50-X)*Hill Slope))
X is the logarithm of concentration. Y is the response.
Y is % stimulation of RFU and starts at Bottom and goes to Top along a sigmoid curve.
Figure 2. Dose dependent stimulation of intracellular cAMP accumulation upon treatment with nicotinic acid in CHO-K1/NIACR1/Gα15 cells. d2 acceptor fluorophore -labeled cAMP (Cat. No. 62AM4PEC; Revvity) and intracellular cAMP in CHO-K1/NIACR1/Gα15 cells competitively bind with Europium Cryptate-labeled anti-cAMP monoclonal antibody. The FRET signal decreases as the intracellular cAMP concentration rises and was measured by plate reader (Pherastar, BMG). The EC50 of nicotinic acid on CHO-K1/NIACR1/Gα15 cells was 10.80 nM.
For research use only. Not intended for human or animal clinical trials, therapeutic or diagnostic use.
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