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| | Dihydrofolate Reductase (DHFR) Inhibition Assay |
| Description: | The assay mixture contained 50 μM Tris-HCl buffer (pH 7.4), 50 μM NADPH, 10 μl DMSO or the same volume of DMSO solution containing the test compounds to a final concentration of 10^-11-10^-5 M, and 10 μl of bovine liver DHFR, in a final volume of 1.0 ml [Pignatello et al., Pharm. Pharmacol. Commun., 5:299-304]. After addition of the enzyme, the mixture was incubated at room temperature for 2.0 min, and the reaction was initiated by adding 5 μl of dihydrofolic acid, the change in absorbance (ΔOD/min) was measured by the spectrophotometer at 340 nm and 22 °C, kinetic program (reading every 15 s for 2.5 min). |
| Affinity data for this assay | |
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