Depletion of Fetal and Postnatal Macrophages

To deplete YS-derived macrophages pregnant female mice were treated with a single injection of CSF-1R blocking antibody (clone AFS98, Bio X Cell) or rat IgG2a control antibody (clone 2A3, Bio X Cell). Three mg of the antibodies in sterile PBS were administered i.p. to pregnant females at E6.5, as described^{12 (link),32 (link)}. Mice were sacrificed at E17.5 or at postnatal age of 2 wk or 5 wk for flow cytometric analyses.
To deplete tissue-resident macrophages after birth, 2 wk old C57Bl/6N mice were cyclically treated with anti-CSF1 antibody and clodronate (Fig. 2b). To that end, three doses of CSF1 neutralizing antibody (Clone 5A1, BioXcell) or control IgG (clone HRPN, BioXcell) were given i.p. (0.5 mg on postnatal day 14, 0.25 mg on postnatal day 18 and 0.25 mg on postnatal day 22). On subsequent days, three doses of clodronate or control liposomes (Liposoma) 50 µl/injection on postnatal days 15, 19 and 23) were administered i.v. The mice were sacrificed 1 or 11 days after the final clodronate treatment. In control experiments using kidney, we saw a full recovery of a known bone marrow-derived CD11b⁺F4/80^Int macrophage population, but not that of a known fetal-derived CD11b^IntF4/80^Hi macrophage population, verifying the robustness of the model.