Rifampin was incorporated into Middlebrook 7H10 medium either by adding an aliquot of the antibiotic into melted agar (final concentration 1.0 and 3.0 μg/ml) or by submerging a 5 or 15 μg of rifampin paper disk into 5 ml of melted agar contained in one quadrant of a Felson “X” plate. At intervals, plugs of agar were removed from the stored plates and assayed. Plates stored at 5 C for 28 days showed no loss of potency; at 37 C, the half-life of rifampin was 9 days. Stability of rifampin at these concentrations in 7H10 medium was independent of the method used for incorporation. Using the disk method, uniform rifampin concentrations of 0.75 μg/ml on day 5 for the 5-μg disk and 2.7 μg/ml on day 6 for the 15-μg disk were observed. Results indicated that the rifampin concentrations within the agar dilution and disk diffusion plates were equivalent at these times.