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CD8+ T-cell responses

YM Yuki Mizumoto
HH Hiroaki Hemmi
MK Masahiro Katsuda
MM Motoki Miyazawa
YK Yuji Kitahata
AM Atsushi Miyamoto
MN Mikihito Nakamori
TO Toshiyasu Ojima
KM Kenji Matsuda
MN Masaki Nakamura
KH Keiji Hayata
YF Yuri Fukuda-Ohta
MS Masanaka Sugiyama
TO Tomokazu Ohta
TO Takashi Orimo
SO Soichiro Okura
IS Izumi Sasaki
KT Koji Tamada
HY Hiroki Yamaue
TK Tsuneyasu Kaisho
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Mice were immunised with the indicated doses of OT-I peptides, XCL1-OT-I or OVA protein with or without 20 µg of poly(I:C). At 7 days after immunisation, splenocytes were prepared and subjected to FACS. To detect OT-I Ag-specific CD8+ T cells, splenocytes were stained with a H-2Kb/OVA tetramer (MBL) and mAbs for CD8α, CD49b and CD62L. For intracellular IFN-γ staining, splenocytes were stimulated with 1 µg/ml of OT-I peptides for 6 h in the presence of brefeldin A (10 µg/ml, Sigma-Aldrich), and stained with mAbs for CD8α, CD49b and CD62L. Cells were further fixed with fixation/permeabilisation solution (Cytofix/ Cytoperm Kit, BD Bioscience), and stained with anti-IFN-γ mAb. Percentages of tetramer-positive or IFN-γ+ cells among CD8α+CD49b−CD62L− T cells were calculated.

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