Human LPMC [lamina propria mononuclear cells] were isolated as previously described.58 (link) Murine LPMC were isolated as in Burrello et al.23 (link) Briefly, the dissected human intestinal mucosa was freed of mucus and epithelial cells in sequential steps with DTT [0.1 mmol/L] and EDTA [1 mmol/L] [both from Sigma] and then digested with collagenase D [400 U/ml] [Worthington Biochemical Corporation, Lakewood, NJ] for 5 h at 37°. LPMCs were then separated with a Percoll gradient and cultured in complete RPMI 1640 medium containing 5% human serum [Sigma] and 100 UI/ml IL-2 [Proleukin].
For murine LPMC isolation, colonic lamina propria mononuclear cells were isolated via incubation with 5 mM EDTA at 37°C for 30 min, followed by mechanical disruption with GentleMACS [Miltenyi Biotec]. After filtration with 100-µm and 70-µm nylon strainers [BD], the LPMC were counted and stained for immunophenotyping.
For murine LPMC isolation, colonic lamina propria mononuclear cells were isolated via incubation with 5 mM EDTA at 37°C for 30 min, followed by mechanical disruption with GentleMACS [Miltenyi Biotec]. After filtration with 100-µm and 70-µm nylon strainers [BD], the LPMC were counted and stained for immunophenotyping.
