During the last decade, protein analysis and proteomics have been established as new tools for understanding various biological problems. As the identification of proteins after classical separation techniques, such as two-dimensional gel electrophoresis, have become standard methods, new challenges arise in the field of proteomics. The development of "functional proteomics" combines functional characterization, like regulation, localization and modification, with the identification of proteins for deeper insight into cellular functions. Therefore, different mass spectrometric techniques for the analysis of post-translational modifications, such as phosphorylation and glycosylation, have been established as well as isolation and separation methods for the analysis of highly complex samples, e.g. protein complexes or cell organelles. Furthermore, quantification of protein levels within cells is becoming a focus of interest as mass spectrometric methods for relative or even absolute quantification have currently not been available. Protein or genome databases have been an essential part of protein identification up to now. Thus, de novo sequencing offers new possibilities in protein analytical studies of organisms not yet completely sequenced. The intention of this review is to provide a short overview about the current capabilities of protein analysis when addressing various biological problems.