The only vaccine currently available for the prevention of tuberculosis in man is a live attenuated vaccine, bacille Calmette-Guerin (BCG), derived from Mycobacterium bovis. Concerns over the lack of the universal efficacy and safety of BCG have resulted in efforts to develop a new generation of TB vaccines. Historically, killed whole-cell preparations of mycobacteria have been ineffective vaccines. We revisited the potential of killed whole-cell vaccines by comparing their efficacy with live BCG Pasteur in a guinea pig challenge model. BCG Pasteur was inactivated with a low concentration of formalin and showed to be non-viable in culture or severe combined immunodeficient mice. Formalin-inactivated BCG was mixed with non-phospholipid liposome adjuvants (Novasomes) and administered to guinea pigs as a single subcutaneous inoculation. All formulations were well tolerated and one conferred a significant survival advantage against lethal aerogenic challenge with M. bovis.