In this publication, the use of ultraviolet (UV) matrix-assisted laser desorption/ionization (MALDI) time-of-fight (TOF) mass spectrometry (MS) for rapid identification and characterization of Saccharomyces cerevisiae, a fungus, is reported. S. cerevisiae is a unicellular eukaroyte that can serve as a model to study more complex organisms. We have determined that the best technique for cell wall lyses for MALDI involves the use of high concentrations of formic acid solutions. We also have shown that different fungal species exhibit different mass spectra, which can be used to distinguish them readily. Protein peaks from S. cerevisiae spectra have been tentatively identified using bioinformatics and are mainly assigned to ribosomal and mitochondrion-related proteins.