CD11b+ pre-DCs were isolated from the BM of isotype or TNFα-depleted infected and control mice by magnetic bead separation as described above according to the manufacturer’s protocol. Cells were washed and counted, and then 20,000 cells were cytospun onto charged glass microscope slides, fixed, stained with primary antibodies against H3K4me3 (Active Motif, Carlsbad, CA) and Fizz1 (R&D Systems, Minneapolis, MN) or control immunoglobulin G (IgG) (R&D Systems), then stained using anti-rabbit Alexa Fluor 594 or anti-goat Alexa Fluor 488 secondary antibodies, and mounted with VECTASHIELD mounting medium plus 4′,6-diamidino-2-phenylindole (DAPI) (Vector Laboratories, Burlingame, CA). Slides were visualized by confocal microscopy using a spinning disk confocal microscope (Olympus America Inc., Center Valley, PA) with a digital charge-coupled device camera (Hamamatsu Photonics, Hamamatsu, Japan) for image capture and an arc lamp illumination source providing excitation wavelengths of 350 to 700 nm and three-color emission analyses. The acquired digital images were processed and analyzed using Stereo Investigator software version 9 (MBF Bioscience, Williston, VT).

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