To date, the diagnosis of tuberculosis has not improved significantly and still relies heavily on staining and culture of sputum or other clinical specimens which were developed more than 100 years ago. Staining does not differentiate tuberculosis from other mycobacterial infections, and culture requires at least 4 to 8 weeks. These are the major problems faced by tuberculosis control programs. In response to this demand, new rapid diagnostic methods are urgently sought. In recent years, much hope has been laid on the development of molecular techniques in the routine tuberculosis laboratory. This review concentrates on 4 techniques that are increasingly used in clinical laboratories: polymerase chain reaction to detect mycobacterial DNA in clinical specimens, nucleic acid probes to identify culture, restriction fragment length polymorphism analysis to compare strains for epidemiologic purposes, and genetic-base susceptibility testing methods for rapid detection of drug resistance. Finally, the increase in the use of clinically-useful molecular biological techniques that affect turnaround time, length of stay, and patient outcome, and reduce overall hospitalization costs will continue until universal standardization for molecular diagnostic procedures are provided. At present, conventional methods should not be replaced by novel methods until the latter are shown to be of equal or greater sensitivity, specificity, reliability, and user-friendliness. However, it is expected that the newly developed molecular techniques will complement our armamentarium of diagnostic tools in the detection of tuberculosis. It is also expected that clinical protocols based on molecular methods will increase the chances for cure by selecting the most appropriate treatment and improving the quality of life of tuberculosis patients.