Comprehensive Immunophenotyping of Cells

Cells were fixed in 2% paraformaldehyde (PFA; Electron Microscopy Sciences), permeabilized using the BD Cytofix/Cytoperm kit when observing intracellular proteins, and stained with primary antibodies followed by species-matching secondary antibodies (Invitrogen). Primary antibodies were: anti-CCR5 (Santa Cruz), anti-SAMHD1 (polyclonal antibody; Sigma), anti-CD82 (TS82b, kindly provided by Dr. E. Rubinstein, Villejuif, France), and antibodies produced in our laboratory (anti-CD9 (VJ1/20), anti-CD151 (LIA1/1), anti-CD63 (Tea3/18), and anti-CD4 (HP2/6))17 (link), 49 (link), 50 (link). Data was acquired with a FACSCantoII flow cytometer (BD), and analysed with BD FACSDIVA (BD) or FlowJo (FlowJo LLC) softwares.

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Rocha-Perugini V., Suárez H., Álvarez S., López-Martín S., Lenzi G.M., Vences-Catalán F., Levy S., Kim B., Muñoz-Fernández M.A., Sánchez-Madrid F, & Yáñez-Mó M. (2017). CD81 association with SAMHD1 enhances HIV-1 reverse transcription by increasing dNTP levels. Nature microbiology, 2(11), 1513-1522.

Publication 2017

PFA BD Cytofix/Cytoperm kit species-matching secondary antibodies anti-CCR5 anti-SAMHD1 anti-CD82 ( FACSCantoII flow cytometer BD FACSDIVA

Antibodies Antibody Ccr5 Cd151 Cells Electron microscopy Intracellular Paraformaldehyde Proteins Samhd1

Corresponding Organization :

Other organizations : Hospital Universitario de La Princesa, Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid, Hospital General Universitario Gregorio Marañón, Emory University, Stanford University, Centro de Investigación en Red en Enfermedades Cardiovasculares

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Variable analysis

independent variables

Cell permeabilization using the BD Cytofix/Cytoperm kit when observing intracellular proteins
Primary antibody staining: anti-CCR5, anti-SAMHD1, anti-CD82, anti-CD9, anti-CD151, anti-CD63, anti-CD4

dependent variables

Expression of cell surface proteins: CCR5, CD82, CD9, CD151, CD63, CD4

control variables

Cell fixation in 2% paraformaldehyde (PFA)
Use of species-matching secondary antibodies (Invitrogen)

positive controls

Antibodies produced in the authors' laboratory (anti-CD9 (VJ1/20), anti-CD151 (LIA1/1), anti-CD63 (Tea3/18), and anti-CD4 (HP2/6))

negative controls

Not explicitly mentioned

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