Abscess and empyema fluid supernatants have zinc-reversible antimicrobial activity that is thought to be due to calprotectin, a calcium- and zinc-binding protein present within neutrophil cytoplasm. The present studies were undertaken to determine if calprotectin isolated from human empyema fluid supernatants demonstrated similar antimicrobial activity to that of the original specimens. The characteristics of the calprotectin complex on SDS-PAGE and Western blotting with specific antisera were similar in neutrophil lysates and in empyema fluid supernatants. Ion-exchange and size-exclusion chromatography were used to obtain highly purified preparations of calprotectin from empyema fluids, and these preparations demonstrated zinc-reversible anti-Candida albicans activity which was similar to that observed in the original specimens. These findings suggest that calprotectin is responsible for most of the growth-inhibitory activity of empyema fluid supernatants against this organism.