An in vitro system to investigate the ability of macrophages to recognize and ingest senescent polymorphonuclear neutrophils has been used that uses chromium-labeled neutrophils and staining for myeloperoxidase (MPO). Human monocyte-derived macrophages obtained from in vitro cultures were able to recognize "aged" but not freshly isolated 51Cr-labeled human neutrophils and ingest them. Freshly isolated monocytes did not exhibit this property. Because the aged neutrophils were greater than 95% viable, death did not appear to be a prerequisite for recognition and ingestion. Serum was not required for the aging of the neutrophils, and when serum was used, different concentrations did not appear to effect the aging process; that is, neutrophils aged in different concentrations of serum were ingested equally. Phagocytosis of senescent neutrophils by macrophages occurred in a time-dependent manner and was also dependent on the number of neutrophils added. Monocyte-derived macrophages first exhibited the ability to phagocytose senescent neutrophils on the 3rd d of culture, with the percentage of active macrophages increasing through day 7. In experiments with rabbit mononuclear phagocytes, immune complex-induced inflammatory macrophages from the lung but not resident bronchoalveolar macrophages or peripheral blood monocytes were found to be capable of recognition and ingestion of senescent rabbit neutrophils. These data suggest that the monocyte maturation process, akin to that seen during inflammation, is necessary in vitro before macrophages recognize and remove senescent neutrophils.