DAT Human Dopamine Transporter Functional Antagonist Uptake LeadHunter Assay - TW
OVERVIEW
DOCUMENTS/REFERENCES
Biological Information
Background Information:
The actions of several neurotransmitters, including dopamine, are regulated through their rapid uptake and clearance from synaptic junctions by plasma membrane transport proteins. The dopamine transporter in central dopaminergic neurones is responsible for the recovery of up to 90% of released transmitters. The monoamine transporters are high-affinity targets for a number of psychoactive agents such as cocaine, amphetamine, and antidepressants. These agents, by blocking transporters and consequently preventing neuronal uptake, elevate levels of extracellular neurotransmitter concentrations in both the central and peripheral nervous system, contributing to their behavioral and autonomic effects.
Family:
Transporter
Sub Family:
SLC6 / Monoamine
Class:
SLC
Protein Name:
DAT
Uniprot Number:
Q01959
Protein Aliases:
dopamine transporter
Gene Name:
SLC6A3
Gene ID:
6531
Gene Aliases:
DAT|DAT1
Accession Number:
NM_001044
Organism:
Human
Construct Details:
Full Length, Recombinant
Cell Type:
CHO-S
Assay Information
Assay Type:
Functional
Assay Sub Type:
Cell Based
Functional Mode:
Antagonist
Detection Method:
Radiometry
Measured Response:
Scintillation
Testing Information
Procedure Summary:
Human recombinant dopamine transporter expressed in CHO-S/hDAT clone 28 cells are plated. Test compound and/or vehicle is preincubated with cells (1 x 10⁵/ml) in modified Tris-HEPES buffer pH 7.1 for 20 minutes at 25℃ and 50 nM [³H]Dopamine is then added for an additional 10 minute incubation period. A lysate is obtained from solubilized cells and counted to determine [³H]Dopamine uptake. Reduction of [³H]Dopamine uptake by 50 percent or more (≥50%) relative to 10 µM nomifensine indicates significant inhibitory activity. If significant inhibition of uptake is observed, item #316110-1 is suggested as follow up assay to evaluate for possible compound-induced cytotoxicity.
Incubation:
10 min at 25°C
Control Inhibitor:
Nomifensine
Criteria For Significance:
≥ 50% Inhibition of [3H]Dopamine uptake relative to nomifensine response
Test Concentration / Dose:
Compounds are screened at 10, 3, 1, 0.3, 0.1, 0.03, 0.01 and 0.003 μM
Test Sample Requirements:
75 μl of 10 mM stock or 1 mg (pre-weighed) powder (assuming FW ≦500 and purity 100%)
Minimum Order Quantity:
16
Turnaround Time
Standard:
20 Business Days
Reference Compound Data
Clinical Relevance
Therapeutic Area:
CNS/Addiction/Pain
Additional Information
Brand:
LeadHunter
Testing Location:
Taiwan - Taipei
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