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determine whether macrophages from Ptdsr-knockout mice were impaired in the efficacy of apoptotic cell uptake in vitro, we performed phagocytosis assays with fetal-liver-derived macrophages (FLDMs) and quantified their phagocytosis rates. Phagocytosis of apoptotic thymocytes was investigated at 60, 90 and 120 minutes after addition of target cells in the absence of serum. Analysis of phagocytosis rates by flow cytometric analysis (FACS) revealed no differences in the efficacy of apoptotic cell uptake between Ptdsr -/- and Ptdsr+/+ macrophages and demonstrated no differences in apoptotic cell engulfment between selected time points (data not shown). To re-examine and further independently validate the result of normal apoptotic cell uptake by Ptdsr -/- macrophages, we performed phagocytosis assays for 60 min and determined the percentage of macrophages that had engulfed apoptotic cells, in a total of at least 300 macrophages counted by fluorescence microscopy. Phagocytosed, 5-carboxytetramethylrhodamine- (TAMRA-) labeled apoptotic cells were identified as being engulfed by inclusion in F4/80-labeled macrophages. Analysis was done independently by three investigators who were not aware of macrophage genotypes (Ptdsr -/- or Ptdsr+/+). Again, no differences were found in the percentage of macrophages that had engulfed apoptotic cells (Figure 7a,c,e) or in the relative number of phagocytosed apoptotic cells per macrophage (phagocytotic index; Figure 7f). Moreover, single Ptdsr -/-macrophages could be identified that had engulfed even more apoptotic target cells than had wild-type macrophages (Figure 7b,d). Thus, Ptdsr-deficient macrophages had a normal ability to ingest apoptotic cells and were not impaired in recognition or phagocytosis of cells that had undergone programmed cell death."}

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determine whether macrophages from Ptdsr-knockout mice were impaired in the efficacy of apoptotic cell uptake in vitro, we performed phagocytosis assays with fetal-liver-derived macrophages (FLDMs) and quantified their phagocytosis rates. Phagocytosis of apoptotic thymocytes was investigated at 60, 90 and 120 minutes after addition of target cells in the absence of serum. Analysis of phagocytosis rates by flow cytometric analysis (FACS) revealed no differences in the efficacy of apoptotic cell uptake between Ptdsr -/- and Ptdsr+/+ macrophages and demonstrated no differences in apoptotic cell engulfment between selected time points (data not shown). To re-examine and further independently validate the result of normal apoptotic cell uptake by Ptdsr -/- macrophages, we performed phagocytosis assays for 60 min and determined the percentage of macrophages that had engulfed apoptotic cells, in a total of at least 300 macrophages counted by fluorescence microscopy. Phagocytosed, 5-carboxytetramethylrhodamine- (TAMRA-) labeled apoptotic cells were identified as being engulfed by inclusion in F4/80-labeled macrophages. Analysis was done independently by three investigators who were not aware of macrophage genotypes (Ptdsr -/- or Ptdsr+/+). Again, no differences were found in the percentage of macrophages that had engulfed apoptotic cells (Figure 7a,c,e) or in the relative number of phagocytosed apoptotic cells per macrophage (phagocytotic index; Figure 7f). Moreover, single Ptdsr -/-macrophages could be identified that had engulfed even more apoptotic target cells than had wild-type macrophages (Figure 7b,d). Thus, Ptdsr-deficient macrophages had a normal ability to ingest apoptotic cells and were not impaired in recognition or phagocytosis of cells that had undergone programmed cell death."}

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Phagocytosis of apoptotic thymocytes was investigated at 60, 90 and 120 minutes after addition of target cells in the absence of serum. Analysis of phagocytosis rates by flow cytometric analysis (FACS) revealed no differences in the efficacy of apoptotic cell uptake between Ptdsr -/- and Ptdsr+/+ macrophages and demonstrated no differences in apoptotic cell engulfment between selected time points (data not shown). To re-examine and further independently validate the result of normal apoptotic cell uptake by Ptdsr -/- macrophages, we performed phagocytosis assays for 60 min and determined the percentage of macrophages that had engulfed apoptotic cells, in a total of at least 300 macrophages counted by fluorescence microscopy. Phagocytosed, 5-carboxytetramethylrhodamine- (TAMRA-) labeled apoptotic cells were identified as being engulfed by inclusion in F4/80-labeled macrophages. Analysis was done independently by three investigators who were not aware of macrophage genotypes (Ptdsr -/- or Ptdsr+/+). Again, no differences were found in the percentage of macrophages that had engulfed apoptotic cells (Figure 7a,c,e) or in the relative number of phagocytosed apoptotic cells per macrophage (phagocytotic index; Figure 7f). Moreover, single Ptdsr -/-macrophages could be identified that had engulfed even more apoptotic target cells than had wild-type macrophages (Figure 7b,d). Thus, Ptdsr-deficient macrophages had a normal ability to ingest apoptotic cells and were not impaired in recognition or phagocytosis of cells that had undergone programmed cell death."}