BACKGROUND: Various combinations of creatine kinase-MB, myoglobin, and cardiac troponin I or T (cTnI/cTnT) have been used to evaluate patients with suspected acute coronary syndromes. The current recommendation is to use the 99th percentile of cTnI/cTnT as the sole marker for diagnosis of acute myocardial infarction. METHODS AND RESULTS: We retrospectively analyzed cardiac marker protocols collected from 824 US hospitals undergoing Chest Pain Center Accreditation through the Society of Cardiovascular Patient Care from 2009 to 2014. Data were obtained by a self-reported survey that addressed cardiac marker(s), sampling time periods, and cut points used for evaluation of suspected acute myocardial infarction. The combination of cTnI or cTnT with creatine kinase-MB was the most commonly used biomarker strategy. Use of cTnI or cTnT as the sole marker increased over time (14-37%; P<0.0001), as did use of the 99th percentile cut point for cTnI/cTnT (30-60%; P<0.0001). CONCLUSION: There is considerable variation in cardiac marker testing strategies used in US hospitals for evaluation of suspected acute myocardial infarction. Although increasing, 24% of hospitals used a cTn alone strategy, and only 49% used cTn at the recommended 99th percentile cut point. This has important implications for the diagnosis and treatment of patients with acute myocardial infarction.