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Product Images
Figure 1. Calcium flux in A1-expressing Chem-3 cell line induced by N6-cyclopentyladenosine (CPA). A1-expressing Chem-3 cells ...
Figure 2. Assay for antagonist activity at A1 by calcium flux assay. A1-expressing Chem-3 cells were loaded with Fluo-4 and washed. DPCPX was added ...
ChemiScreen™ A1 Calcium-Optimized FLIPR Cell Lines
Description:
ChemiScreen™ A1 Calcium-Optimized FLIPR Cell Lines
Trade Name:
ChemiScreen
Product Overview:
TRANSFECTION: Full-length human ADORA1 cDNA encoding A1
Background Information:
Extracellular adenosine mediates a multitude of biological effects, including wakefulness, antiarrythmia, bronchoconstriction and response to ischemia and oxidative stress. A family of four GPCR adenosine receptors, A1, A2A, A2B and A3, is responsible for these effects. The A1 receptor, which couples to Gi/o, is most highly expressed in brain, and mediates endogenous antinociception and neuronal response to hypoxia (Fredholm et al., 2001). A1 is also expressed in kidney, where it contributes to tubuloglomerular feedback. Chemicon's cloned human A1-expressing cell line is made in the Chem-3 host, which supports high levels of recombinant A1 expression on the cell surface and contains high levels of the promiscuous G protein Gα15 to couple the receptor to the calcium signaling pathway. Thus, the cell line is an ideal tool for screening for antagonists of interactions between A1 and its ligands.
Application Notes:
Calcium flux assay, ligand binding assays
Species Reactivity:
Human
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Cell Line Type:
GPCR Cell Lines
Concentration:
2 x 106 cells/mL
Host Cells:
Chem-3, a suspension cell line endogenously expressing the promiscuous G protein, Gα15, and lacking endogenous expression of adenosine receptors.
Packaging:
1 mL per vial
Materials Required but Not Delivered:
GROWTH MEDIA: RPMI containing 4.5 g/L glucose/10% heat inactivated fetal bovine serum/0.8 mg/ml Geneticin (G418)/100 U/ml each penicillin and streptomycin, containing 1:100 dilution of Chem-3 Growth Supplement (Chemicon catalog # HTSCHEM-3S).
Cell Type:
Chem-3
Gene Symbol:
- ADORA1
- RDC7
Format:
Cell Line
Presentation:
Cells are frozen at 2 x 106 cells/mL in RPMI/20% fetal bovine serum/100 U/ml penicillin and streptomycin/10% DMSO. Cell line tests negative for mycoplasma.
Quality Assurance:
SPECIFICATIONS:
EC50 for calcium mobilization by CPA: ~ 0.8 nM
IC50 for DPCPX: ~250 nM
Signal:noise ratio at CPA Emax: 747
EC50 for calcium mobilization by CPA: ~ 0.8 nM
IC50 for DPCPX: ~250 nM
Signal:noise ratio at CPA Emax: 747
Protein or Enzyme Type:
GPCRs
Species:
Human
Storage Conditions:
Place cells in liquid nitrogen immediately upon receipt. Maintain frozen in liquid nitrogen for up to 5 years. Thaw cells rapidly by removing from liquid nitrogen and immediately immersing in a 37ºC water bath. Immediately after ice has thawed, sterilize the exterior of the vial with 70% ethanol. Transfer contents of the vial to a sterile conical tube containing 10 mL growth media, and centrifuge at 200 x g. Remove supernatant and resuspend in fresh growth media. Transfer to a T75 flask containing 20 mL final volume growth media. Cells should be maintained between 0.1 x 106 and 1.0 x 106 cells/mL, and are typically passaged 1:10 every 3-4 days. Cells should be passaged at least once after thawing prior to use in calcium flux assays.