Isolation and Purification of CD27+ Microparticles

For proliferation assays, CD27⁺ MPs were sorted with a MoFlo Astrios flow cytometer (Beckman Coulter, Brea, CA) equipped with a PMT-FSC detector, as previously described (13 (link), 15 (link)). Flow cytometer performance was assessed with Megamix-Plus FSC and SSC beads (BioCytex) before the MP sorting experiments. MPs were labeled with anti-CD27-BV510 (BD Biosciences) antibody (4°C, 30 minutes). We purified CD27⁺ MPs with diameters between 200 and 900 nm. We used a commercial kit to check for the absence of endotoxin in the sorted CD27⁺ MP preparations (In vivogen, San Diego, CA).

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Cagnet L., Neyrinck-Leglantier D., Tamagne M., Berradhia L., Khelfa M., Cleophax S., Pirenne F, & Vingert B. (2023). CD27+ microparticle interactions and immunoregulation of CD4+ T lymphocytes. Frontiers in Immunology, 14, 1043255.

Publication 2023

MoFlo Astrios flow cytometer Megamix-Plus FSC anti-CD27-BV510

Antibody Assays Endotoxin

Corresponding Organization :

Other organizations : Université Paris-Est Créteil, Laboratory of Excellence GR-Ex, Établissement Français du Sang, Inserm, Institut Mondor de Recherche Biomédicale

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Variable analysis

independent variables

Sorting CD27+ MPs with a MoFlo Astrios flow cytometer

dependent variables

Proliferation of CD27+ MPs

control variables

Assessing flow cytometer performance with Megamix-Plus FSC and SSC beads before MP sorting
Labeling CD27+ MPs with anti-CD27-BV510 antibody
Purifying CD27+ MPs with diameters between 200 and 900 nm
Checking for the absence of endotoxin in the sorted CD27+ MP preparations using a commercial kit

controls

Positive control: Not specified
Negative control: Not specified

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