Intrapleural tissue plasminogen activator (tPA) and deoxyribonuclease (DNase) therapy is being increasingly employed as an alternative to surgical intervention for the treatment of complicated parapneumonic effusions and empyema. Published cases are limited to one randomized control trial and few case reports. No data exist on employing sequential or repeated courses of intrapleural tPA/DNase to aid evacuation of separate collections in patients' with a multiloculated pleural infection. This is the first report of successful use of sequential delivery of separate courses of intrapleural tPA/DNase to two noncommunicating infected pleural fluid collections within the same hemithorax of a patient. Our case confirms that prior treatment with tPA/DNase therapy does not preclude subsequent effective and safe use of this intrapleural treatment.