Omental Stromal Cell Immunostaining

Omenta were fixed in 2% neutral buffered formalin for 1 h on ice, before staining in the fridge overnight with antibodies to GATA6 (Cell Signalling; clone D61E4; purified) and F4/80 (Serotec; clone CL:A3-1; purified) or TIM4 (Biolegend; clone RMT4-54; conjugated to AF647) in PBS containing 0.5% BSA, 0.5% triton X-100 (Sigma). Omenta were subsequently stained for 1 h at room temperature with indicated antibodies (Table S2) in PBS containing 0.5% BSA, 0.5% triton X-100, with DAPI added for the final 10 min. Samples were mounted with Fluoromount G (ThermoFischer) and confocal images acquired using a Leica SP8 laser scanning confocal microscope using Leica LAS X software. 3D reconstruction was created using LAS-X-3D (Leica) v3.5.7.23225. Gata6 expression in CD45⁻ omental cells was determined by analysis of a previously pubished dataset available under accession number GSM4053741 [23 (link)].