The effect of four media or media combinations on the growth and the in vitro susceptibility to amphotericin B, 5-fluorocytosine, fluconazole and itraconazole of 51 isolates of Cr. neoformans was studied. The drug concentration producing a 50% decrease in OD405 (IC50) was determined in microtiter format. The following MOPS buffered media or media combinations (antifungal drug diluent/inoculum diluent) were used: RPMI 1640 (RPMI)/RPMI, Yeast nitrogen base (YNB)/YNB, RPMI/YNB and RPMI/High resolution broth. Amphotericin B was further tested in M3 broth in the microdilution format and on RPMI agar with the E-test. Growth rates (OD405) were higher in YNB, RPMI/YNB, RPMI/HR and M3 as compared to RPMI alone. After 48 h of incubation, IC50 values obtained with RPMI alone and with RPMI/YNB correlated within two dilutions for 98, 95, 95 and 100% of test strains with amphotericin B, 5-fluorocytosine,fluconazole and itraconazole respectively. The E-test provided the widest range of IC values with amphotericin B. Growth of Cr. neoformans can be enhanced in susceptibility tests using microtiter plates with drugs diluted in RPMI by preparing the inoculum in YNB. However, when using spectrophotometric endpoint determinations, sufficient growth is obtainable with most isolates for test interpretation at 48 h in RPMI alone.