Acute myocardial infarction (AMI) patients are at an elevated risk for life-threatening myocardial ischemia/reperfusion injury. Early-stage nonradioactive and noninvasive diagnosis of AMI is imperative for the subsequent disease treatment, yet it presents substantial challenges. After AMI, the myocardium typically exhibits elevated levels of peroxynitrite (ONOO−), constituting a distinct microenvironmental feature. In this context, the near-infrared imaging probe (BBEB) is employed to precisely delineate the boundaries of AMI lesions with a high level of sensitivity and specificity by monitoring endogenous ONOO−. This probe allows for the early detection of myocardial damage at cellular and animal levels, providing exceptional temporal and spatial resolution. Notably, BBEB enables visualization of ONOO− level alterations during AMI treatment incorporating antioxidant drugs. Overall, BBEB can rapidly and accurately visualize myocardial injury, particularly in the early stages, and can further facilitate antioxidant drug screening.