Fig. 2: Tumor-infiltrating myeloid immune cells and dendritic cells (DCs) display the highest levels of TGFβ isoform expression.

B16 tumors from mice were harvested 11 days following tumor implantation. After creating single-cell suspensions, tumors underwent processing and staining for flow cytometry analysis as described in “Methods”. a Breakdown of the immune infiltrate in B16 tumors of mice harvested 11 days after tumor implantation. b Representative histograms displaying mean fluorescence intensity (MFI) of either TGFβ1 (left panel) or TGFβ3 (right panel) on specific tumor-infiltrating immune cells as detected by flow cytometry. The light gray peak represents each cell type’s fluorescence minus one (FMO) and was used to determine positive expression, indicated by the colored peak. c Representative graph illustrating the relative expression of TGFβ1 (top graph) or TGFβ3 (bottom graph) by various lymphocytic and myeloid cell types in the tumor microenvironment 11 days after tumor implantation. Data (n = 5 mice/group) are displayed as MFI \(\pm\) SD. Data are representative of three independent experiments. MFI is measured in arbitrary units and is a variable used to measure relative expression levels of staining antibodies, in this case of TGFβ1 and TGFβ3 protein expression, on tumor-infiltrating immune cells. FMO controls were derived by staining the immune cells with all the fluorophores minus one fluorophore, in this case, the fluorophore (Alexa Fluor 647) that was conjugated to TGFβ1 and TGFβ3. The pattern of intracellular TGFβ1 and TGFβ3 expression is similar to the surface staining pattern demonstrated in Supplementary Fig. 3b.