A single cell suspension from spleens was prepared by passing through a nylon 40 mm cell strainer (BD, San Jose, CA). Suspensions were washed with RPMI complete (10% FCS, 1%Pennicilin/Streptomycin, 1% Glutamine, 1% Sodium Pyruvate, 1% nonessential amino acids, 0.1% B-mercaptoethanol) (Life Technologies, Grand Island, NY) and centrifuged for 5 minutes at 1200 RPM at 4uC. Red Blood Cells were lysed using 0.86% ammonium chloride solution, centrifuged and resuspended in RPMI complete. BMNCs were prepared as previously described [89] .