H1 Human Histamine GPCR Binding Antagonist Radioligand LeadHunter Assay - TW

Procedure Summary:

Human recombinant histamine H1 receptor expressed in CHO-K1 cells are used in modified Tris-HCl buffer pH 7.4. A 10 μg‡ aliquot is incubated with 1.2 nM [³H]Pyrilamine for 180 minutes at 25℃. Non-specific binding is estimated in the presence of 1 μM pyrilamine. Receptor proteins are filtered and washed, the filters are then counted to determine [³H]Pyrilamine 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] Pyrilamine

Ligand Kd (nM):

1.1

Ligand Concentration:

1.2 nM

Non Specific:

1 µM Pyrilamine

Incubation:

180 min at 25°C

Control Inhibitor:

Pyrilamine

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