Fungi growing on building materials represent an exposure risk for atopic patients and various methods of control have been proposed. Recent studies have investigated the in vitro antifungal activity of Melaleuca alternifolia (tea tree) oil against fungal pathogens. The goal of this study was to investigate the effect of tea tree oil, (TTO), on fungi normally found growing on building materials. Ceiling tile and sheet rock squares (36 cm³) were sterilized and soaked with sterile distilled water (SDW), various concentrations of TTO or a commercial mold inhibitor, (MI). Materials were inoculated with a spore suspension of one of the following fungi: Aspergillus niger, Aspergillus versicolor, Stachybotrys chartarum, Alternaria sp., Cladosporium sp., and Penicillium sp. Cultures were incubated at room temperature for up to 12 weeks. Fungal growth was assessed by measuring colony diameter starting at week one. Spray applications of TTO were also investigated. TTO inhibited all six fungal isolates to some degree when grown on ceiling tile and sheet rock. Of the various concentrations 1.0% TTO was the only concentration that exhibited total growth inhibition of all six isolates. The MI inhibited growth for four of the six fungi tested but did not inhibit the growth of Alternaria or Stachybotrys. TTO spray applications in 1.0% concentration did not completely inhibit fungal growth; however, 5.0% TTO spray application successfully inhibited all growth. TTO, in all concentrations demonstrated antifungal activity on sheet rock and ceiling tile building materials; therefore showing possible applications for control of indoor fungal contaminants.