added to each sample and incubated for an additional 3 hours at 37uC, 5% CO2 in RPMI containing 10% Human Serum. Samples were washed once with 1 ml PBS containing 10% FBS and stained for surface antigens using CD8 PerCP-Cy5.5 or CD4 PerCP- Cy5.5 antibodies at 4uC in the dark for 20 minutes. After wash with PBS containing 2% FBS and re-suspension in 150 ml of 2% BD Cytofix cells were incubated at room temperature in the dark for 20 minutes. Samples were washed twice with 1 ml of Perm/ Wash buffer (BD Bioscience) and incubated at room temperature in the dark for 20 minutes with 2 ml of purified Mouse IgG1 antibody. Samples were stained for intra-cellular cytokines using IL-2 PE and IFN-c APC antibodies at room temperature in the dark for 20 minutes. Samples were washed twice with 1 ml of BD Perm/Wash buffer and re-suspended in 150 ml of 2% BD Cytofix, acquired on a BD FACSCalibur flow cytometer and analyzed using FlowJo software. PBMC that had proliferated (CFSE ‘low’) REFERENCES 1. 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