The role of biochemical markers in the diagnosis of acute coronary syndromes has increased considerably in the past decade. The World Health Organization previously defined acute myocardial infarction as a combination of at least 2 of 3 components: symptoms consistent with acute myocardial infarction, electrocardiogram changes diagnostic of acute myocardial infarction, and an enzyme pattern with classic rise and fall. Measurement of creatine kinase and its MB fraction by various assays was the gold standard for the diagnosis. Troponins are more specific and sensitive markers for myocardial injury, and their increasing utilization has resulted in a broadening of the definition of acute myocardial infarction to incorporate high-risk acute coronary syndromes. Previously, traditional enzyme evaluation left patients with small amounts of cellular death undiagnosed; these patients were categorized as having unstable angina or, worse, noncardiac chest pain. Newer markers now identify these patients as a subgroup at high risk for cardiac death or cardiac events. Newer therapeutic interventions and a more invasive strategy have been shown to improve outcomes in this high-risk subgroup. Increased specificity has also reduced the number of patients who undergo extensive, expensive, and invasive evaluations for noncardiac syndromes due to false elevations of traditional markers. This article comprehensively reviews the evolution of biochemical markers for the diagnosis of acute myocardial infarction, addressing their promise for improving delivery of care and outcomes and their technical and diagnostic pitfalls.