| Product Description |
Recombinant CHO-K1 cells stably overexpress human opioid receptor kappa 1 (OPRK1) on the surface and contain high levels of G protein Gαi to couple with the receptor in downstream signaling pathways. |
| Applications |
Calcium flux assay, IP-One assay and cAMP accumulation assay. |
| Expressed Gene |
NCBI reference sequence NM_000912; no expressed tags |
| Target Protein |
NP_000903 |
| 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, Life Technologies), 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, Life Technologies) |
| Gene Synonyms |
K-OR-1, KOP, KOR, KOR-1, KOR1, OPRK |
| Background |
Opioid receptors and their endogenous peptide ligands play important roles in the reward and reinforcement of drugs such as heroin, cocaine, and alcohol. The κ-opioid receptor is a type of opioid receptor which binds the peptide opioid dynorphin as the primary endogenous ligand. κ-opioid receptors are widely distributed in the brain, spinal cord, and in pain neurons. They are associated with the risk for alcohol dependence. |
Figure 2. Dose dependent stimulation of intracellular cAMP accumulation upon treatment with U-50488 in CHO-K1/OPRK1/Gα15 cells. d2 acceptor fluorophore -labeled cAMP (Cat. No. 62AM4PEC; Revvity) and intracellular cAMP in CHO-K1/OPRK1/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 U-50488 on CHO-K1/OPRK1/Gα15 cells was 42.36 nM.
Figure 1. U-50488-induced concentration-dependent stimulation of intracellular calcium mobilization in CHO-K1/OPRK1/Gα15 cells. The cells were loaded with Calcium-4 (Cat. No. R8142; Molecular Devices) prior to stimulation with OPRK1 agonist, U-50488. 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 U-50488 (mean ± SEM, n = 3). The EC50 of U-50488 on CHO-K1/OPRK1/Gα15 cells was 11.07 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 RFU and starts at Bottom and goes to Top along a sigmoid curve.
For research use only. Not intended for human or animal clinical trials, therapeutic or diagnostic use.
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