Fungi were isolated from various cultural heritage objects and morphologically identified. Fungi, moulds in particular, are the most common causes of cultural heritage deterioration, and their toxicity and pathogenicity may be hazardous to human health. A total of 24 samples were sampled from 12 cultural heritage objects, at four locations in Istria. Substrates selected for sampling were fresco, paper, wood, textile and glass. Fungi were cultured on Malt Extract Agar (MEA) and Czapek Dox Agar (CHD) and their growth was monitored. Morphological characteristics were studied by using light and scanning electron microscopy. A total of 21 different groups were isolated and grouped into six genera. Of these, five were determined to species level. The following genera were identified: Aspergillus, Aureobasidium, Chaetomium, Epicoccum, Penicillium and Stachybotrys. For a more accurate determination and results verification molecular analyses are required. The results will be used for the development of laboratory protocol for the isolation and morphological identification of fungi from cultural heritage objects for the restorers’, conservators’ and archaeologists’ requirements as well as for review catalog for the Material Research Center METRIS and other users. Thus the results and extensive photographic documentation will get practical application.