For immune-monitoring studies, patient PBMC samples were stained with cocktail fluorescent-conjugated antibody mix diluted in PBS containing 2% FBS + 0.01% azide for 30 min at 4 °C in the dark. After a 2-step washing with PBS, cells were fixed with 2% PFA for 20 min at 4 °C. Samples were analyzed on a BD Fortessa Flow Cytometer (UV-Violet-Blue-Yellow/Green-Red 5-Laser configuration). For the first cohort (aviremic patients) the following antibodies were used: CCR5-FITC, CD158a-PE, CD57-PE-CF594, CD14-Alexa Fluor 700, CD19-Alexa Fluor 700, CXCR4-BV421, CD62L-BV605, NKG2D-BV650, CD16-BV711, CD3-BV786, CD56-BUV395 and CD4-BUV737 (BD Biosciences), CD158b-PE, CD158e/k-PE, NKp46-PE-Vio770, NKG2C-APC, CD45RA-APC-Vio770, and CD45RO-VioGreen (Miltenyi). For the second cohort (viremic patients) the similar panel of antibodies were re-used, except for additional antibodies for CXCR4-BV421 and CD107a-PE-CF594 (BD Biosciences). Cell death and viability were determined by DAPI (BD Biosciences). Data were analyzed using FlowJo software (v10.8) (BD Biosciences) with appropriate plugins for high-dimensional analysis and visualization.
Vo D.N., Leventoux N., Campos-Mora M., Gimenez S., Corbeau P, & Villalba M. (2022). NK Cells Acquire CCR5 and CXCR4 by Trogocytosis in People Living with HIV-1. Vaccines, 10(5), 688.
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