Anti-BrdU (3D4), anti-Ly5.2 (104), anti-Thy1.1 (OX-7), anti-CD62L
(MEL-14), anti-IFNγ (XMG1.2), anti-TNF (MP6-XT2) were from BD
Biosciences; anti-CD8α (53–6.7), anti-KLRG-1 (2F1), anti-IL-2
(JE56–5H4), anti-CD44 (IM7), Bcl-2 (633504), anti-mouse Perforin Antibody
(S16009A), anti-human/mouse granzyme B Antibody (GB11) were from Biolegend;
anti-TCF1 (C63D9) was from Cell Signaling Technology. For intracellular staining
of Tcf1, Bcl-2, granzyme B and perforin, cells were fixed and permeabilized
(eBioscience, 00–5524). Leukocyte Activation Cocktail containing phorbol
myristate acetate (PMA) and ionomycin (BD Biosciences) was used to stimulate T
cells for intracellular cytokine staining. A Fixation/Permeabilization Solution
Kit (BD Biosciences) was used to fix and permeabilize the cells. Annexin V
staining was performed with Annexin V Apoptosis Detection Kit (eBiosciences).
BrdU staining was performed with BrdU Staining Kit (eBiosciences) following the
protocol provided by the manufacturer. LSR II or BDFortessa (BD Biosciences)
were used for flow cytometry acquisition. Samples were analyzed with FlowJo
software (TreeStar). Naive CD8+ T cells were enriched using
Naïve CD8+ T cell isolation kit from Stem Cell Technology. A
FACSAria (BD Biosciences) was employed for all other T cell enrichments.
(MEL-14), anti-IFNγ (XMG1.2), anti-TNF (MP6-XT2) were from BD
Biosciences; anti-CD8α (53–6.7), anti-KLRG-1 (2F1), anti-IL-2
(JE56–5H4), anti-CD44 (IM7), Bcl-2 (633504), anti-mouse Perforin Antibody
(S16009A), anti-human/mouse granzyme B Antibody (GB11) were from Biolegend;
anti-TCF1 (C63D9) was from Cell Signaling Technology. For intracellular staining
of Tcf1, Bcl-2, granzyme B and perforin, cells were fixed and permeabilized
(eBioscience, 00–5524). Leukocyte Activation Cocktail containing phorbol
myristate acetate (PMA) and ionomycin (BD Biosciences) was used to stimulate T
cells for intracellular cytokine staining. A Fixation/Permeabilization Solution
Kit (BD Biosciences) was used to fix and permeabilize the cells. Annexin V
staining was performed with Annexin V Apoptosis Detection Kit (eBiosciences).
BrdU staining was performed with BrdU Staining Kit (eBiosciences) following the
protocol provided by the manufacturer. LSR II or BDFortessa (BD Biosciences)
were used for flow cytometry acquisition. Samples were analyzed with FlowJo
software (TreeStar). Naive CD8+ T cells were enriched using
Naïve CD8+ T cell isolation kit from Stem Cell Technology. A
FACSAria (BD Biosciences) was employed for all other T cell enrichments.