All organisms contain thousands of proteins and peptides in their body fluids. A deeper insight into the functional relevance of these polypeptides under different physiological and pathophysiological conditions and the discovery of specific peptide biomarkers would greatly enhance diagnosis and therapy of specific diseases. The low-molecular-weight proteome, also termed peptidome, provides a rich source of information. Due to its unique features, the technical challenges differ somewhat from those in "common" proteomics. In this paper, we focus on the low-molecular-weight urinary proteome. We review the methodological aspects of sample collection, preparation, analysis, and subsequent data evaluation. In the second part of this review, we summarize the recent progress in the definition and identification of clinically relevant polypeptide markers. © 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.