The aims of this study were to evaluate free levels of fluconazole (FCZ) in the kidneys of healthy and Candida albicansinfected Wistar rats using microdialysis and to establish the relationship between free renal and total plasma levels under both conditions. Microdialysis recovery rates were determined in vitro by dialysis, and retrodialysis recovery rates were determined in vivo by retrodialysis. The recovery rate was around 50%, independent of the method, drug concentration, or condition (in vitro or in vivo) used. FCZ kidney penetration in healthy and infected rats was investigated after the administration of 10 mg/kg of body weight intravenously (i.v.) or 50 mg/kg orally (n ؍ 6/group) and blood and microdialysate sample harvesting at predetermined time points up to 24 and 18 h, respectively. There were no statistical differences between the area under the free concentration-time curve (AUC 0 -ؕ ) values in plasma and in tissue for either healthy or infected groups for the same dose regimen investigated. The antifungal tissue penetrations were similar for both doses and under all conditions investigated (ranging from 0.77 to 0.84). The unbound fraction of FCZ was concentration independent (86.0% ؎ 2.0%), allowing the prediction of free renal levels using pharmacokinetic parameters obtained from total plasma fitting. The results showed that free renal and free plasma levels are similar in healthy and systemically C. albicansinfected rats. Therefore, free plasma levels are a good surrogate to estimate free FCZ renal concentrations in systemic candidiasis and can be used to optimize dosing regimens for this drug.