The objective was to determine if an intra-articular injection of an extracellular matrix (ECM) powder and blood composite (ECM-B) after anterior cruciate ligament (ACL) injury would have a mitigating effect on posttraumatic osteoarthritis and if that effect would be different with terminal sterilization of the ECM powder before use. Eighty Lewis rats underwent ACL transection and were divided into four groups:​ (1) intra-articular injection with phosphate-buffered saline (PBS;​ n = 20), (2) intra-articular injection of ECM-B using aseptically processed ECM (ASEPTIC;​ n = 20), (3) intra-articular injection of the ECM-busing ECM powder sterilized with 15 kGy electron beam irradiation (EBEAM;​ n = 20), and (4) intra-articular injection of the ECM-B using ECM powder sterilized with ethylene oxide (EO;​ n = 20). Twenty additional animals received capsulotomy only (SHAM). The animals were followed for 6 weeks and evaluations of gait, radiographs, and joint cartilage histology were performed. At 6 weeks, when compared to the SHAM group, the group treated with PBS had significantly worse gait and histologic changes, while the ASEPTIC group was not different from SHAM for either of these outcomes. When compared to the SHAM group, the EO group had similar gait outcomes, but greater histologic damage, and the EBEAM group had significantly worse gait and histological outcomes. The ECM-B composite produced using aseptically processed ECM powder mitigated the gait and histologic changes associated with osteoarthritis after ACL transection in the rat;​ however, care must be taken when selecting a terminal sterilization method as this may affect the effectiveness of treatment.