Traditional drug susceptibility methods can take several days or weeks of incubation between drug exposure and confirmation of sensitivity or resistance. In addition, these methods do not capture information about viable organisms that are not immediately culturable after drug exposure. Here, we present a rapid fluorescence detection method for assessing the susceptibility of M. tuberculosis to antibiotics. Fluorescent markers Calcein violet-AM and SYTOX-green are used for measuring cell viability or cell death and to capture information about the susceptibility of the whole population and not just those bacteria that can grow in media postexposure. Postexposure to the antibiotic, the method gives a rapid readout of drug susceptibility, as well as insights into the concentration and time-dependent drug activity following antibiotic exposure.