Listeria Monocytogenes Infection Model

L. monocytogenes of the wild-type, attenuated (LADD), or Δhly background were grown overnight in brain heart infusion media at 30C. The bacteria were back diluted 1:5 and allowed to grow to log phase (OD0.4–0.6, ~1–1.5 hours) at 37C, with aeration, prior to infection. Bacteria were diluted in PBS and mice were infected with 200μL at the indicated doses intravenously. For bacterial burden analysis, mice were sacrificed at 12hpi and livers were homogenized in 0.1% Nonidet P-40 in PBS and plated on Luria-Bertani plates. For splenic macrophage depletion, 200μL clodronate, PBS control, or endosome lipid control (Encapsula Nano Sciences) were given intravenously 24 hours prior to bacterial infection according to the manufacturer’s instructions. For B-cell depletion, mice were dosed intraperitoneally with 12.5μg of Ultra-LEAF Purified anti-mouse CD20 (clone SA271G2) or isotype control (clone RTK4530) in 100μL PBS 24 hours prior to bacterial infection. Depletion efficacy of relevant cell subsets was confirmed by assessing abundance of splenic CD11b⁺ cells (clone M1/70), CD11c⁺ cells (clone N418), or B220⁺ cells (clone RA3-6B2) by flow cytometry. Celecoxib (Cayman Chemical) was milled into standard mouse chow (Envigo) at 100mg/kg and fed ad lib for 48 hours before and after immunization [16 (link),77 (link)].