VMAT (Non-Selective) Human Vesicular Monoamine Transporter Binding LeadHunter Assay - TW

Procedure Summary:

This assay measures binding of [³H]DHTB to sites associated with the vesicular monoamine uptake transporter. Human concentrated platelets from Chinese Blood Services Foundation obtained are prepared in modified HEPES buffer pH 8.0. A 0.175 mg‡ aliquot of membrane protein is incubated with 2 nM [³H]DHTB for 60 minutes at 25℃. Non-specific binding is estimated in the presence of 10 μM Ro4-1284. Membranes are filtered and washed, the filters are then counted to determine [³H]DHTB specifically bound. Compounds are screened at 10 μM.
Note: ‡Membrane protein may change from lot to lot, the concentration used will be adjusted if necessary.

Ligand:

[³H] Dihydrotetrabenazine

Ligand Kd (nM):

2.5

Ligand Concentration:

2 nM

Non Specific:

10 µM Ro4-1284

Incubation:

60 min at 25°C

Control Inhibitor:

Tetrabenazine

Criteria For Significance:

≥ 50% of max stimulation or inhibition

Test Concentration / Dose:

Suggested concentration for initial testing is 10 uM

Test Sample Requirements:

Minimum for 1) Screen: 25 μl of 10 mM stock -OR- 1 mg (pre-weighed). 2) Dose Response: 60 μl of 10 mM stock -OR- 1 mg (pre-weighed).

Minimum Order Quantity:

2