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Fig. 3 - Supplemental 3 Simultaneous inactivation of Pu.1 and Spi-b leads to rapid elimination of microglia in zebrafish. (A) The experimental setup for pu.1 conditional knockout in adult spi-bΔ232/Δ232 mutants and the representative images of the midbrain cross section of adult pu.1KI/+;spi-bΔ232/Δ232;Tg(coro1a:CreER) and pu.1KI/Δ839;spi-bΔ232/Δ232;Tg(coro1a:CreER) fish at 2 dpi and 8 dpi. The white arrow indicates microglia with blebbing morphology. (B–C) Quantification of the number (B) and percentage (C) of DsRed+ microglia in adult pu.1KI/+;spi-bΔ232/Δ232;Tg(coro1a:CreER) and pu.1KI/Δ839;spi-bΔ232/Δ232;Tg(coro1a:CreER) fish at 2 and 8 dpi (n=3 for all groups). (D) The experimental setup for pu.1 conditional knockout in spi-bΔ232/Δ232 mutant embryos and the representative images show the DsRed +microglia in pu.1KI/+;spi-bΔ232/Δ232;Tg(coro1a:CreER) and pu.1KI/Δ839;spi-bΔ232/Δ232;Tg(coro1a:CreER) embryos at 3 and 5 dpf. (E) Quantification of the number of DsRed +microglia in pu.1KI/+;spi-bΔ232/Δ232;Tg(coro1a:CreER) and pu.1KI/Δ839;spi-bΔ232/Δ232;Tg(coro1a:CreER) embryos at 3 and 5 dpf after 4-OHT treatment from 48 to 60 hpf. (pu.1KI/+;spi-bΔ232/Δ232;Tg(coro1a:CreER) n=4 and 6 for 3 and 5 dpf, respectively. pu.1KI/Δ839;spi-bΔ232/Δ232;Tg(coro1a:CreER) n=8 for both 3 and 5 dpf) n.s.=not significant, p>0.05;***p<0.001;****p<0.0001.