Computational protocol: IFNγ Regulates Activated Vδ2+ T Cells through a Feedback Mechanism Mediated by Mesenchymal Stem Cells

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Protocol publication

[…] Antibody targets and fluorochromes were as follows: CD4-PerCP-eFluor® 710 (clone SK3), CD8-PerCP-eFluor® 710 (clone SK1), CD3-PE-Cy7 (clone UCHT1), CD45RA-APC-eFluor® 780 (clone HI100) and CD27-APC (clone LG.7F9), all from eBioscience (San Diego, CA, USA). Vδ2 TCR-PE (clone B6) and IFNγ-FITC for intracellular staining (clone 4S.B3) were from BD Pharmingen™ (San Diego, CA, USA). Intracellular staining was done with BD Cytofix/Cytoperm™ Fixation/Permeabilization Solution Kit with BD GolgiStop™Cells (BD Cytofix/Cytoperm™ Plus). For CFSE labeling of Vδ2+ cells the Cell Trace CFSE Cell Proliferation Kit (Invitrogen, San Diego, CA, USA) was used. AnnexinV-DY634 for detection of apoptotic cell death in MSCs was from Immunostep. Surface marker expression, intracellular cytokine production, cell proliferation and AnnexinV staining were analysed on a FACSCanto (BD Biosciences, Chicago, IL, USA) using BD FACSDiva™ software for acquisition. Analysis of flow cytometry data, including CFSE tracking assays and Proliferation Index [] were done with FlowJo software v9.5.3 (TreeStar, Ashland, OR, USA). All staining, CFSE labeling, and AnnexinV staining, were performed according to the manufacturer’s protocol. […]

Pipeline specifications

Software tools BD FACSDiva, FlowJo
Application Flow cytometry
Organisms Homo sapiens
Diseases Autoimmune Diseases, Neoplasms
Chemicals Amino Acids