Isolation and Characterization of CD4+ T Cells

Participant recruitment and informed consent were performed under Institutional Review Board (IRB)-approved protocols at the US National Institutes of Health (NIH) and University of Washington. Peripheral blood mononuclear cells (PBMC) were isolated from whole blood by density gradient centrifugation. PBMC were incubated with Fcγ-receptor-blocking reagent for 10 minutes and stained with LIVE/DEAD Aqua stain, CD3-APC-H7 (BD Biosciences; Cat# 641406), CD4-BV785 (BioLegend; Cat# 317442), CD8-PacBlue (Invitrogen; Cat# MHCD0828), CD14-BV650 (BioLegend; Cat# 301836), CD16-PerCP/Cy5.5 (BioLegend; Cat# 302028), CD19-BV605 (BD Biosciences; Cat# 562653), CD20-BV570 (BioLegend; Cat# 302332), CD27-Alx700 (BioLegend; Cat# 302814), CD32-PE (BioLegend; Cat# 303206); CD45RO-ECD (Beckman Coulter; Cat# IM2712U), CD123-PE/Cy5 (BD Biosciences; Cat# 551065), and TCRγδ-APC (BD Biosciences; Cat# 555718). CD4+ T cells were isolated by fluorescence-activated cell sorting (FACS) on a FACSAria (Becton Dickinson) using previously described protocols^{32 (link)}.

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Sun C., Liu L., Pérez L., Li X., Liu Y., Xu P., Boritz E.A., Mullins J.I, & Abate A.R. (2022). Droplet microfluidic sequencing of HIV proviruses with their integration sites in patients receiving antiretroviral therapy. Nature biomedical engineering, 6(8), 1004-1012.

Publication 2022

CD4-BV785 CD8-PacBlue CD14-BV650 CD16-PerCP/Cy5.5 CD19-BV605 CD20-BV570 CD27-Alx700 CD32-PE CD45RO-ECD CD123-PE/Cy5 TCRγδ-APC FACSAria

Blood Cd123 Cd4 t cells Cd45ro Cy5 5 Density gradient centrifugation Fcγ receptor Institutional review board Peripheral blood mononuclear cells Stain Tcrγδ

Corresponding Organization : QB3

Other organizations : National Institute of Allergy and Infectious Diseases, National Institutes of Health, University of Washington

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Variable analysis

independent variables

Fcγ-receptor-blocking reagent incubation time (10 minutes)

dependent variables

CD3-APC-H7 expression
CD4-BV785 expression
CD8-PacBlue expression
CD14-BV650 expression
CD16-PerCP/Cy5.5 expression
CD19-BV605 expression
CD20-BV570 expression
CD27-Alx700 expression
CD32-PE expression
CD45RO-ECD expression
CD123-PE/Cy5 expression
TCRγδ-APC expression

control variables

Participant recruitment and informed consent procedures under Institutional Review Board (IRB)-approved protocols
Peripheral blood mononuclear cells (PBMC) isolation by density gradient centrifugation
LIVE/DEAD Aqua staining

Annotations

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