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Assay Method Information

Assay Name:  Inhibitor Screening Using Bacterial System (IC50) and Measurement of Inhibition Constant (Ki)
Description:  Nineteen Pyr analogs were studied for their inhibition activity against cells expressing either WT or SP21 mutant PvDHFR-TS. The assays were conducted with 96-well microplates by monitoring the growth at an optical density of 595 nm (A595). The average A595 of control culture omitting Pyr analogs was scored as 100% growth, and the average readings for the cultures at each drug concentration were divided by this value to obtain relative growth values. The concentrations that inhibited 50% bacterial growth (IC50s) were determined from dose-response curves. Inhibition constants (Ki) of antifolates were calculated using a nonlinear least square fit of the data to the Michaelis-Menten equation, assuming the inhibitor binds competitively to the enzyme active site.
Affinity data for this assay
 

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Last update November 1, 2007
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