The burgeoning field of proteomics plays a powerful and relevant role in the discovery of biomarkers, which are biometric measurements that convey information about the biological condition of the subject being tested. Biomarkers have changed the manner in which we diagnose disease, monitor the effect of therapy, classify disease, detect toxicity, and develop new drugs. The central part that proteins command in both disease etiology and treatment make them prime biomarker candidates. Indeed, the majority of clinical tests in use today measure proteins. This perspective introduces the Journal of Proteome Research Special Issue on Proteomics and Biomarkers. It outlines the major applications of biomarkers, discusses the basics of statistically assessing them and considers the crucial choice of sample type. Central considerations of biomarker discovery and validation, particularly with respect to their intended clinical and research applications, are highlighted.