Anti cd4 and anti cd8 antibodies

PBMC were cultured in the presence of media or VSV-G-pseudotyped HIV-1 in the absence or presence of described concentrations of AZT or EFV or Raltegavir for 24 hours. Afterwards, BDCA1⁺ cDCs were purified by immunomagnetic enrichment and mixed with allogeneic total peripheral blood T lymphocytes previously stained with 5μM carboxyfluorescein succinimidyl ester (CFSE; Invitrogen) at a T:DC ratio of 4:1. As a control, T cells were also cultured in the presence of media only or 2.5μg/ml PHA and 50IU/ml IL-2. After incubation for 6 days, cells were washed, stained with viability dye and anti-CD4 and anti-CD8 antibodies (Biolegend, San Diego, CA), and CFSE dilution on CD4 and CD8 T cell subpopulations was analyzed by flow cytometry using a Fortessa flow cytometer.

FACS purified viable CD64^Hi,PD-L1^Hi and CD64^Lo,PD-L1^Lo mDC subpopulations, generated after 24 h of infection with a VSV-G HIV-1 virus or 24 h of incubation with TLR3 ligands (2 μg/mL poly I:C and nanoparticle-loaded gag-dsDNA adjuvants), were mixed with allogeneic total peripheral blood T lymphocytes previously stained with 5 μM carboxyfluorescein succinimidyl ester (CFSE, Invitrogen) at a T:DC ratio of 4:1. As a control, T cells were also cultured in the presence of media alone or 2.5 μg/mL PHA (Sigma) and 50 IU/mL IL-2 (NIH AIDS reagent program). After incubation for six days, cells were washed, stained with viability dye and anti-CD4 and anti-CD8 antibodies (BioLegend, San Diego, CA, USA), and CFSE dilution on CD4⁺ and CD8⁺ T cell subpopulations was analyzed by flow cytometry using a Fortessa flow cytometer.