Isolation of Peripheral Blood Leukocytes

The isolation of peripheral blood leukocytes (PBL) was done as previously described^{14 (link),17 (link)} with methodology similar to one previously described³⁰ . Whole blood was incubated in the same proportion with red blood lysis buffer solution [162.64 mM NH₄Cl (Sigma-Aldrich), 9.98 mM Tris base (Merck, Darmstadt, Germany), pH = 7.2] for 10 min, with agitation, at room temperature. Cells were then passed through a 100-μm cell strainer, washed, and resuspended in complete RPMI medium: RPMI 1640 Medium supplemented with 10% foetal bovine serum (FBS) (FBS South America Premium, Ref. S181BH-500 from Biowest, Nuaillé, France), 85 units/mL penicillin, 85 μg/mL streptomycin, 62.5 ng/mL amphotericin B, 0.05 mM 2-mercaptoethanol and 10 mM HEPES (all from Sigma-Aldrich) after centrifugation at 300×g for 5 min.