Although proteomic technologies have been enthusiastically embraced in the field of biomarker discovery and particularly with biological fluids, it is increasingly being recognized that pre‐analytical effects, i.e. those occurring prior to the point of actual sample analysis and including factors such as sample processing and storage, can exert marked influences on the results obtained. Such effects have been recognized already for specific analytes in clinical chemistry, but with the increasing sensitivity and resolution of the newer technologies, such effects are potentially even more marked. The challenge of translating initial findings into clinical application requires such issues to be addressed at a very early stage in study design and this paper reviews the current knowledge of the potential impact of pre‐analytical factors on proteomic studies of biological fluids.