Ingested RBCs in splenic macrophages of untreated (but not clodronate-treated) mice.

Ingested RBCs are readily apparent in splenic macrophages of untreated mice (but not in clodronate-treated mice). Mice were injected with antibody-opsonized RBCs (10⁹). At 4 to 8 hours later, they were killed and the splenocytes were dispersed, stained, and cytospun onto slides. RBCs were lysed prior to staining until no free RBCs were visible on the slides. (A) Cells were stained for F4/80, a macrophage-specific cell surface molecule, (in green) then fixed and permeabilized for intracellular staining with Ter-119, an RBC specific antibody (in red). (B) Cells were intracellularly stained with anti-CD68 (in red) and anti-RBC (Ter-119, in green). (C) Cells were stained intracellularly for CD68 (in red), and RBCs (in green) are visualized because they were labeled with CFSE prior to administration. In all images, nuclei were counterstained in blue. No ingested RBCs were visible in mice pretreated with liposomal clodronate (data not shown). Original magnifications × 100.