H 2kb siinfekl complex

Cells were stained with combinations of fluorophore-conjugated antibodies against CD11b, CD11c, CD69, CD8, H-2K^b, H-2K^b-SIINFEKL complex (Biolegend), Gr-1 and IFN-γ (BD). For intracellular staining of IFN-γ, cells were incubated for 4 h at the presence of Leukocyte Activation Cocktail (BD), fixed with 4% paraformaldehyde and permeabilized with BD Perm/Wash buffer, followed by IFN-γ antibody staining. For H-2K^b-SIINFEKL Tetramer staining, cells were incubated at 4°C for 1 h. Alexa 488-labeled H-2K^b - SIINFEKL Tetramer was kindly provided by the NIH Tetramer Core at Emory University (Atlanta, GA, USA). Data were collected with a CyAn flow cytometer (Dako) or an Accuri C6 flow cytometer (BD) and analyzed with FlowJo software (Tree Star). Gating strategies were shown in Supporting Information Fig. 8.