Eleven isolates of Cryptococcus neoformans were investigated to determine the biochemical basis of their tolerance to fluconazole. The MICs of fluconazole for three isolates with low-level resistance were 3- to 6-fold higher than those for sensitive isolates, while the MICs for four isolates with high-level resistance were 100- to 200-fold higher than those for sensitive isolates. The level of ergosterol present in the isolates varied, and those which had relatively low levels of ergosterol were resistant to amphotericin B. Changes in the affinity of the target enzyme (sterol 14alpha-demethylase) and decreases in the cellular content of fluconazole seemed to be responsible for the resistance in isolates with low-level and high-level resistance, respectively.