animal models of myocardial ischemia-reperfusion (I/R) mimic the clinical scenario where a period of ischemia is followed by the restoration of myocardial blood flow (e.g., angioplasty or thrombolysis for treatment of acute myocardial infarction) and are important tools for testing novel strategies aimed at reducing ischemic damage and attenuating reperfusion injury. The mouse is increasingly the species of choice for in vivo I/R experiments, because of its genetic malleability, rapid breeding cycle, and economic husbandry. However, the small size of the mouse anatomy imposes challenges not only for surgery itself but also for subsequent histology, with the measurement of infarct size as the prime scientific readout. Unlike after permanent occlusion of a coronary artery where there is necrosis of the entire coronary territory, infarction after I/R is variable, nontransmural, and usually incomplete. Therefore, the principal requirement of I/R histology is the clear demarcation of three myocardial subsets: 1) the unaffected, remote myocardium; 2) the area at risk (AAR), i.e., the myocardium distal to the site of coronary occlusion that was initially subjected to ischemia; and 3) the area of necrosis (AON), which is the fraction of nonviable tissue within the AAR. The final infarct size is expressed as the ratio of AON to AAR, and to this end, gross histological double-staining techniques have served as a gold standard in many studies (e.g., see Refs. 2, 5, 8, 12).