Despite similarities of cellular membranes in all eukaryotes, every compartment displays characteristic and oftenunique features which are important for the functions of the specific organelles. In the present study, we bio-chemically characterized the plasma membrane of the methylotrophic yeastPichia pastoriswith emphasis onthe lipids which form the matrixof this compartment. Prerequisite for this effort was the design of a standardizedand reliable isolation protocol of the plasma membrane at high purity. Analysis of isolated plasma membranesamples fromP. pastorisrevealed an increase of phosphatidylserine and a decrease of phosphatidylcholine com-pared to bulk membranes. The amount of saturated fatty acids in the plasma membrane was higher than in totalcell extracts. Ergosterol, thefinal product of the yeast sterol biosynthetic pathway, was found to be enriched inplasma membrane fractions, although markedly lower than inSaccharomyces cerevisiae. A further characteristicfeature of the plasma membrane fromP. pastoriswas the enrichment of inositol phosphorylceramides over neu-tral sphingolipids, which accumulated in internal membranes. The detailed analysis of theP. pastorisplasmamembrane is discussed in the light of cell biological features of this microorganism especially as a microbialcell factory for heterologous protein production