Celllytic mt solution

0.5–1 x 10⁶ ileum and colon cells from each mouse were stained in FACS buffer for 30 min at 4° C with the following antibodies: CD3-PerCP-Cy5.5 (17A2, Biolegend), CD4-FITC (RM4-4, eBioscience), CD8-BV510 (53–6.7, Biolegend), CD69-PE-Cy7 (h1.2F3, eBioscience), CD103-PE-Dazzle-594 (2E7, Biolegend). Mesenteric lymph node cells were additionally stained with CD44-AlexaFluor-700 (IM7, eBioscience), and CD62L-APC-eFluor780 (MEL-14, eBioscience). Cells were then washed with FACS buffer and fixed in 1% paraformaldehyde before being acquired on a BD LSRii flow cytometer. Colon samples that had less than 1,000 T cells were excluded. Plasma samples were thawed and diluted 1:10 before being assayed for sCD14 by ELISA (R&D Systems) according to the manufacturer’s instructions. Ileum and colon tissues cultured for 24 h at 37° C, 5% CO₂ in complete RPMI were digested with CellLytic MT solution with proteinase inhibitor (SIGMA), and assayed for myeloperoxidase concentrations by ELISA (R&D Systems). ELISA plates were read using a Vmax Kinetic plate reader (Molecular Devices). Tissue sections from the distal end of the ileum and proximal end of the colon were processed for histology and H&E stained. Scoring [34 (link)] was performed by a pathologist blinded to experimental conditions.