Anti p24 kc57 rd1

2 x 10⁵ FACS-sorted Siglec-9⁺ and Siglec-9^- NK cells were cocultured with autologous HIV-infected CD4⁺ T cells (2 x 10⁴ cells/well) for 16 h at 37°C. Following incubation, cells were harvested and evaluated for intracellular p24 by staining with Zombie NIR fixable viability dye (BioLegend), anti-CD3 BV421 (clone UCHT1) (BioLegend), and anti-p24 KC57-RD1 (Beckman Coulter). For Siglec-9 blocking experiments, 2 x 10⁵ purified NK cells in 100 μl medium were treated with anti-Siglec-9 antibody (100 μg/ml) or isotype-matched control antibody for 30 min at 37°C in 96-well microplates before co-culture with autologous HIV-infected CD4⁺ T cells (2 x 10⁴ cells/well) for 16 h at 37°C.

FDCs were fixed in 1% paraformaldehyde solution. Fixed samples were pre-incubated with anti- FcR (2.4G2, house produced) and 2% bovine serum albumin for blocking nonspecific and Fc-mediated binding. Anti-PE (Rockland), donkey anti-rabbit DyLight 405, anti-CD35, anti-Lamp-1 (Biolegend), fluorescent transferrin and anti-p24 (KC57-RD1, Beckman-Coulter) were used for staining ex vivo cultures of FDCs. Images were acquired with a FluoView FV1000 confocal microscope (Olympus) with a 20x lens (NA: 0.7) or a 60x water immersion lens (NA: 1.2) and processed with FluoView software (Olympus). Data were analyzed with Fiji software.