Glutamate (NMDA, NR1/NR2A) Human Ion Channel [3H] CGP-39653 Binding LeadHunter Assay - TW

Procedure Summary:

This assay measures the binding of [³H]CGP-39653 to glutamate NR2A receptors. Human recombinant glutamate NR1/NR2A receptors expressed in HEK293 cells are used in Tris-HCl pH 7.4 buffer. A 0.04 mg‡ aliquot of membrane is incubated with 5 nM [³H]CGP-39653 for 120 minutes at 4℃. Non-specific binding is estimated in the presence of 1 mM L-Glutamic Acid. Membranes are filtered and washed 4 times, and the filters are counted to determine [³H]CGP-39653 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] CGP-39653

Ligand Kd (nM):

9.7

Ligand Concentration:

5 nM

Non Specific:

1000 µM L-Glutamic acid

Incubation:

120 min at 4°C

Control Inhibitor:

L-glutamic acid

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