The DIEP flap is currently considered the gold standard for autologous reconstructive breast surgery. Postoperative flap failure due to microvascular postanastomotic thrombotic occlusion is a rare but severe complication. Alteplase, a thrombolytic agent typically used in the setting of an ischemic stroke, myocardial infarction, or pulmonary embolism, has also been injected into the microcirculation of flaps as a rescue procedure due to imminent flap loss. The purpose of this article is to provide an overview and detailed guidance for such a thrombolytic procedure due to suspected thrombotic microsurgical failure in free flap surgery. We report the case of a 43-year-old woman who underwent unilateral breast reconstruction with a DIEP flap at our department. Approximately 12 hours postoperatively, an arterial inflow problem was suspected and revision surgery was performed. Peripheral flap perfusion remained absent without an obvious cause and distal thrombosis was assumed to be present. Therefore, alteplase was gradually injected into the arterial pedicle in the anterograde direction just distal to the anastomosis while clamping the artery proximally. About 3 hours after selective flap thrombolysis, microcirculation of the flap was successfully restored without complications. Anterograde injection of alteplase can successfully salvage a free flap. To our knowledge, evidence for optimal dosing and delivery of alteplase for the treatment of thrombosed DIEP flaps has not been published to date. Our approach presents a therapeutic option that both maximizes alteplase concentration in the flap and minimizes the dosage required for flap salvage to significantly reduce systemic adverse effects.