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Assay Method Information

Assay Name:  Radioligand Binding Assay
Description:  The compounds of the present invention inhibit the binding of PGD2 to its receptor CRTH2. The inhibitory activity can be investigated by a radioligand binding assay (Sawyer et al., Br. J. Pharmocol 2002, 137, 1163-72). The radioligand binding assay was performed at room temperature in binding buffer (10 mM HEPES/KOH pH 7.4, mM MnCl2, with protease inhibitor cocktail tablets), containing 1.5 nM [3H]PGD2 (Amersham, 156 Cie/mmol), and 10 ug of hCRTH2 HEK293 (EBNA) cell membrane protein in a final volume of 100 ul in 96 well plates (Corning, USA). Non-specific binding was determined in the presence of 1 uM PGD2 (Cayman, USA). Competing pyrazine sulfonamides were diluted in dimethylsulphoxide so that the total volume of dimethylsulfoxide was kept constant at 1% dimethylsulphoxide (Me2SO). 10 ul of the pyrazine sulfonamides were added. Incubation (60 min at room temperature) was terminated by rapid filtration through 96 wells hydrophobic GF/C Unifilter plates.
Affinity data for this assay
 

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Last update November 1, 2007
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