13.00 CZKKč Ells were rinsed once with 2 ml of Macitentan assay medium (MEM containing Shahīd Darah
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Ells were rinsed once with 2 ml of assay medium (MEM containing 20 mM HEPES, pH 7.4) and incubated at 37 for 30 min with 1 ml of assay medium containing 1 mM 1-methyl-3-isobutylxanthine in the absence or presence of 50 M forskolin. The cells were lysed with 1 ml 5 trichloroacetic acid with 1 mM ATP to terminate the reaction and were stored at 4 for 1 h. Intracellular [3H]cAMP was isolated by sequential chromatography as described previously [64]. The level of [3H]cAMP was estimated by determining the ratios of [3H]cAMP to total [3H]ATP and [3H]ADP pools.Kwan et al.
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