Evaluating the Effectiveness of a Test Compound against Mycobacteria in Broth

To carry out a resazurin assay, grow M. tuberculosis in 7H9ADST to mid-log phase. Dilute the culture with 7H9ADST to an OD₆₀₀ of 0.01. Use 7H9ADST to dilute the compounds to two times the testing concentrations, and aliquot 100 microliters of each diluted compound into each well of a clear 96-well plate.

Transfer 100 microliters of the diluted bacterial suspension into each well. Incubate the plates at 37 degrees Celsius in a humidified incubator for 5 days. Dissolve 10 milligrams of resazurin in 100 milliliters of deionized water, and filter-sterilize the solution. Add 30 microliters of resazurin solution to the wells. Then, incubate the cells for 48 hours.

Bacterial growth is indicated by a color conversion from blue to pink. Carry out quantitative analysis according to the text protocol.