Anti cd4 and anti mhc 2 antibodies

For ex vivo co-culture experiments, DCs were sorted from iliac lymph nodes based on expression of CD11c and MHC II. Cells were blocked for Fc binding then stained with anti-MHC II antibody and anti-CD11c antibody (eBioscience, San Diego, CA). Cells from dLN were sorted into two groups; an MHC II high/CD11c mid population of migratory DCs and MHC II mid/ CD11c + population of non-migratory DCs. All sorting was done on a FACSAria (BD Biosciences, San Jose, CA). CD4 T-cells that were already enriched using the CD4+ T-cell negative selection kit (Stem Cell Technologies, Vancouver, Canada) were also stained using anti-CD4 and anti-MHC II antibodies (eBioscience, San Diego, CA) and sorted into a population of CD4+ and MHC II – cells to ensure that the CD4 T-cells were not contaminated with antigen presenting cells.
For flow cytometry, cells were incubated in fixable viability dye (eBioscience, San Diego, CA), blocked for Fc binding, then stained for surface proteins. Data were analyzed using FlowJo software (Treestar, Ashland, OR). For more details, including antibodies used, see supplemental experimental procedures.