Medulloblastoma is the most frequent malignant brain tumor in children and is considered to be of neuroectodermal origin. Two main representative cell lines, DAOY and D283, are widely used in studies of medulloblastoma. The former shows expression of neuronal and glial elements whereas the latter is assigned to neuronal lineages. We decided to systematically study the proteome of these cell lines in order to find novel and known proteins that could serve as candidate markers or could be of interest as specific antigens for future vaccines. We studied DAOY and D283 by two-dimensional gel electrophoresis with subsequent matrix-assisted laser desorption/ionization identification. A series of identified medulloblastoma proteins were already described in many other malignancies of different origin. An antiapoptotic principle, Ded protein, was observed in both cell lines. Several hypothetical proteins, that were never described at the protein level but only predicted from nucleic acid sequences, could be identified. We conclude that medulloblastoma proteins SYT interacting protein, similar to glucose related protein 58 kDa, hypothetical 37.5 kDa protein, serologically defined colon cancer antigen 10, hepatocellular carcinoma-associated antigen 59, X-ray repair complementing defective repair in CHO 5, hypothetical protein Q96ir7, nit protein 2 and hypothetical protein Q96e67, have been described in a series of other malignancies possibly indicating a role for those in tumor biology and pathomechanisms. The antiapoptotic principle, Ded protein, found in both cell lineages may stand for immortalization but could also determine malignancy per se in medulloblastoma.