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Staining Cells were washed and suspended at a density of 1x106 cells in 100 pL of PBS with 0.1% sodium azide and 5% normal mouse serum (Sigma, USA), and incubated for 10 min on ice; this was followed by surface staining with anti-CD3 mAb and antiCD4 mAb/anti-CD8 mAb for 20 min on ice in the dark. For intracellular cytokine staining, the cells were washed and fixed in 100 pL of fixation buffer (Caltag, USA) containing 4% paraformaldehyde for 20 min on ice. After a wash, cells were suspended in 100 mL of permeabilization buffer containing 0.1% saponin (Caltag, USA) with 5% normal rat serum (Sigma, USA) for 10 min at room temperature and incubated with anti-IL-10 mAb or isotype control IgG2a for 30 min at room temperature in the dark. After a final wash, cells were suspended in PBS with 1% paraformaldehyde (BDH Inc, Canada) and kept in the dark at 4°C until analysis. Flow cytometric analysis A FACScan flow cytometer (Becton Dickinson, USA) was used for analysis. Ten thousand events were acquired in list mode, with debris excluded by the forward scatter threshold. The data were analyzed using CELLQuest software (Becton Dickinson, USA).