Anti cd8 apc cy7 clone sk1

PBMCs from CMV-positive donors were resuspended in RPMI media supplemented with 2% human AB serum (Corning, Celgro), and labeled with 5 μM CFSE (Molecular Probes, Life Technologies). CFSE-labeled PBMCs were inoculated with the recombinant strains of EBOV at an MOI of 2 PFU/cell, with or without simultaneous stimulation with 2 μg/ml of CMV pp65 peptides, SEB treated, or mock treated for 4 hours. PBMCs were washed twice to remove virus inoculum and cultured at a concentration of 2 x 10⁶ cells/ml for 7 days in Advanced RPMI medium supplemented with 10% human AB serum, 2 mM L-glutamine, 200 IU/ml penicillin, and 200 μg/ml streptomycin sulfate (Invitrogen) in 6 well plates. PBMCs were harvested, stained extracellularly with the following antibodies: anti-CD3 PE-Cy7 (clone SK7, BD Biosciences), anti-CD4-PerCP/Cy5.5 (clone SK3, BD Biosciences), anti-CD8 APC-Cy7 (clone SK1, BD Biosciences), and Live/Dead-Far Red Dead cell stain (Molecular Probes, Life Technologies). Cells were subsequently washed, fixed with 4% paraformaldehyde and removed from the BSL-4 according to an approved protocol. Cells were washed with PBS and analyzed by flow cytometry on BD LSR II.