The clinical application of in vitro antifungal susceptibility testing has been limited by a lack of reproducibility and uncertain clinical relevance. As a result of several collaborative studies, the National Committee for Clinical Laboratory Standards (NCCLS) has proposed a standardized antifungal susceptibility test method, NCCLS M27-T. More convenient, user-friendly methods (microdilution broth and stable gradient technology) have been evaluated, and the potential for a similar process with a disk diffusion method is apparent. Adaptation of the standard method for susceptibility testing of filamentous fungi appears promising. The existence of a standardized method facilitates meaningful analysis of studies addressing the issue of clinical relevance of antifungal susceptibility testing. Correlation of MICs with clinical response to therapy is beginning to emerge, most notably in relation to fluconazole and itraconazole therapy for oropharyngeal candidiasis associated with infection with the human immunodeficiency virus. This accumulated experience with antifungal susceptibility testing allows us to provide several specific recommendations for antifungal susceptibility testing in the clinical laboratory. Application of this developing technology to new antifungal agents and other disease states will enhance our ability to effectively deal with the emerging problem of fungal infection.