Figure 5. Investigating the maturation phenotype of mononuclear phagocytes derived from collagenase digestion and spontaneous migration.

(A) Blood DCs were compared with skin mononuclear phagocytes isolated by either collagenase digestion or spontaneous migration for their expression of CD80, CD83, and CD86 by flow cytometry. Primary mononuclear phagocytes (blood and skin) were also compared with immature and mature MDDCs (iMDDCs and mMDDCs, respectively). The geometric mean fluorescence intensity (gMFI) for each surface expression minus the isotype plotted with the means ± 95% confidence interval. Statistics were generated with the Mann-Whitney U test. *P < 0.05; **P < 0.01; ***P < 0.001. Statistics are not shown for the comparison between mMDDCs and all other subsets because they were significantly different to all cells, excluding those isolated by spontaneous migration. (B) Cells isolated from collagenase-digested dermis were cultured in RPMI-1640 supplemented with gentamicin at 37°C for up to 24 h. Cells were collected immediately after digestion and at time intervals up to 24 h and were examined for their expression of CD80, CD83, CD86, CD54, and MR by flow cytometry. Displayed are the means ± sd of the expression at each time point relative to time 0. AF, autofluorescent.