Chemoresistance remains an unresolved problem in clinical oncology. Therefore it is important to identify molecular factors that lead to an understanding of the mechanisms of drug resistance in cancer cells. On the protein-expression level, this can be done using proteomics, which has become the focus of significant interest and research over the past decade. We describe an easy and practicable standardized technique that can be used to study global protein expression in chemosensitive and chemoresistant cancer cells to find candidate proteins that are potentially associated with the drug-resistant phenotype. As an example, fractionation of human neuroblastoma cells using two-dimensional polyacrylamide electrophoresis, spot detection, image analysis, and finally protein identification is illustrated.