The causative agents of Building Related Illness are unknown but are thought to originate from chemical reactions and/or microbial growth caused by high water activity. The aim of this study was to apply direct determination of mycoflora and selected mycotoxins in building materials, settled and cultured airborne dust from preschools and daycare centers. The mycoflora on building materials was directly identified by light microscopy, viable airborne mold propagules counted using a Reuter Centrifugal Sampler, and settled dust was collected on surfaces above floor level. Mycotoxin analysis performed using gas-chromatography- and high pressure liquid chromatography- tandem mass spectrometry showed that mycotoxins are produced in hidden mold infested materials in the construction, that airborne mold from such materials have the capacity to produce mycotoxins, and that mycotoxins can be found in settled dust in the breathing zone.