Background: Most patients presenting chest complaints in primary care are referred to secondary care facilities, whereas only a few are diagnosed with acute coronary syndrome (ACS). Objective: The aim is to determine the optimal cut-off value for a point-of-care heart-type fatty acid binding protein (H-FABP) test in patients presenting to the emergency department and to evaluate a possible future role of H-FABP in safely ruling out ACS in primary care. Methods: Serial plasma H-FABP (index test) and high sensitivity troponin T (hs-cTnT) (reference test) were determined in patients with any new-onset chest complaint. In a receiver operating characteristic (ROC) curve, the optimal cut-off value of H-FABP for ACS was determined. Predictive values of H-FABP for ACS were calculated. Results: For 202 consecutive patients (prevalence ACS 59%), the ROC curve based on the results of the fi rst H-FABP was equal to the ROC curve of hs-cTnT (AUC 0.79 versus 0.80). Using a cut-off value of 4.0 ng/ml for H-FABP, sensitivity for ACS of the H-FABP (hs-cTnT) tests was 73.9% (70.6%). Negative predictive value (NPV) of H-FABP for ACS in a population representative for primary care (incidence of ACS 22%) thus could reach 90.8%. Conclusion: In patients presenting chest pain, plasma H-FABP reaches the highest diagnostic value when a cut-off value of 4 ng/ml is used. Diagnostic values of an algorithm combining point-of-care H-FABP measurement and a score of signs and symptoms should be studied in primary care, to learn if such an algorithm could safely reduce referral rate by GPs.