Multicolour flow cytometric analysis was performed using the following fluorochrome-labelled anti-human monoclonal antibodies for measuring lymphocyte Th and Tfh subsets (CD4-APC-Vio770, CD45RA-PE-Vio770, CCR6-APC, CXCR3-VioBright FITC CXCR5 PerCP-Cy5.5, CCR4-PE) all from Miltenyi biotech. Tregs and Tfr were detected using BD Human Regulatory T Cell Cocktail (BD, San Jose, CA, USA), including CD4-FITC (clone SK3), CD25-PECy7 (clone 2A3), CD127 Alexa Fluor® 647 and adding CXCR5 PerCP-Cy5 and CD8 horizon450 (the last two from Biolegend). Bregs were detected using the following fluorochrome-labelled anti-human monoclonal antibodies: CD38-FITC CD1d-PE, CD19-PE-Cy7, CD5-PerCP-CY5.5, CD24-APC-H7 and CD27- BV510, all from BD Biosciences (San Jose, CA, USA).
Cd45ra pe vio770
Manufactured by Miltenyi Biotec
Sourced in Italy, Germany
CD45RA-PE-Vio770 is a fluorochrome-conjugated antibody that binds to the CD45RA antigen, which is expressed on a subset of T cells and B cells. It can be used for the identification and enumeration of these cell populations in flow cytometry applications.
Automatically generated - may contain errors
Lab products found in correlation
Cd4 apc vio770, by Miltenyi Biotec (2 mentions)
Ccr6 apc, by Miltenyi Biotec (2 mentions)
Cxcr3 viobright fitc cxcr5 percp cy5, by Miltenyi Biotec (2 mentions)
Ccr4 pe, by Miltenyi Biotec (2 mentions)
Cxcr5 percp cy5, by BioLegend (2 mentions)
Cd8 horizon450, by BioLegend (2 mentions)
Cd127 alexa fluor 647, by BioLegend (1 mentions)
Cd19 pe cy7, by BD (1 mentions)
Cd5 percp cy5, by BD (1 mentions)
Cd27 bv510, by BD (1 mentions)
Live dead fixable near ir dead cell stain kit, by Thermo Fisher Scientific (1 mentions)
Cd4 percp vio700, by Miltenyi Biotec (1 mentions)
Foxp3 alexa fluor 647, by BD (1 mentions)
Cd137 pe, by Miltenyi Biotec (1 mentions)
Cd3 fitc, by Miltenyi Biotec (1 mentions)
Fcr blocking reagent, by Miltenyi Biotec (1 mentions)
Kaluza software 2, by Beckman Coulter (1 mentions)
Cd3 apc cy7 okt3, by BioLegend (1 mentions)
Ctla 4 bv421, by BD (1 mentions)
Cd3 pe vio770, by Miltenyi Biotec (1 mentions)
5 protocols using cd45ra pe vio770
1
Multicolor Flow Cytometry Analysis of Lymphocyte Subsets
2
Comprehensive Immunophenotyping of Activated T Cells
3
Multicolor Flow Cytometry for Immune Cell Profiling
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Multicolor flow cytometric analysis was performed, and the gating strategy is reported in the Supplementary Materials . The first tube was to detect B cell subsets and included the following mAbs: CD19-PE-Cy7 (SJ25C1), CD24-APC-Cy7 (ML5), CD38-FITC (HB-7), CD5-PerCP-Cy5.5 (L17F12), CD1d-PE (all from BD Biosciences, San Jose, CA, USA), CD45-APC (REA747 Miltenyi Biotec S.r.l., 40133 Bologna, Italy), and CD27-BV510 (M-T271 BioLegend, San Diego, CA, USA).
The second tube was to detect follicular T cells and included mAbs: CD4-APC-Vio770, CD45RA-PE-Vio770, CCR6-APC, CXCR3-VioBright FITC CXCR5 PerCP-Cy5.5, and CCR4-PE, all from Miltenyi Biotec (40133 Bologna, Italy).
The third tube was to phenotype regulatory T cells according to the BD Human Regulatory T Cell Cocktail (CD4-FITC (SK3), CD25-PECy7 (2A3), CD127 Alexa Fluor® 647, BD, San Jose, CA, USA), CXCR5 PerCP-Cy5, CD8 horizon450, and CD3 APC-Cy7 (OKT3) (Biolegend, San Diego, CA, USA).
Gating was carried out using Kaluza software 2.1 (Beckman Coulter, Inc., Danvers, MA, USA): details are shown in theSupplementary Materials . The T and B cell subsets are also described in the Supplementary Materials .
The second tube was to detect follicular T cells and included mAbs: CD4-APC-Vio770, CD45RA-PE-Vio770, CCR6-APC, CXCR3-VioBright FITC CXCR5 PerCP-Cy5.5, and CCR4-PE, all from Miltenyi Biotec (40133 Bologna, Italy).
The third tube was to phenotype regulatory T cells according to the BD Human Regulatory T Cell Cocktail (CD4-FITC (SK3), CD25-PECy7 (2A3), CD127 Alexa Fluor® 647, BD, San Jose, CA, USA), CXCR5 PerCP-Cy5, CD8 horizon450, and CD3 APC-Cy7 (OKT3) (Biolegend, San Diego, CA, USA).
Gating was carried out using Kaluza software 2.1 (Beckman Coulter, Inc., Danvers, MA, USA): details are shown in the
4
Multiparametric Analysis of T-cell Subsets
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Foxp3-APC (ebioscience, clone 236A/E7) and mIgG1 κ APC isotype control (ebioscience, clone P3.6.2.8.1), CD25-PE (Miltenyi), CTLA-4-BV421 (BD Biosciences, clone BNI3) and mIgG2a κ BV421 isotype control (BD Biosciences), IFN-γ-PE and mIgG1 κ PE isotype control (ebioscience), IFN-γ-FITC and mIgG1 κ FITC isotype control (ebioscience), IL-17A eFluor450 and mIgG1 κ eFlour450 isotype control (ebioscience), CD45RA-PE-Vio770 (Miltenyi), CD45RA-FITC (Miltenyi), CD45RO-PE (BD Biosciences), CD4-PerCP (BD Biosciences), CD3-PE-Vio770 (Miltenyi), CD8-eFlour450 (ebioscience), fixable viability dye-eFlour780 (ebioscience), CFSE (Molecular Probes).
5
Multiparametric Flow Cytometry Panel
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The following fluorochrome‐labeled monoclonal anti‐human antibodies were used to identify central and effector cells: CD3‐APC‐Cy7, CD4‐V450 (REA623), CD45RA‐PE‐Vio770 (REA562), CD196 (CCR6)‐APC (REA190), CD183 (CXCR3)‐VioBright FITC (REA232), CD194 (CCR4)‐PE (REA279) and PerCP‐Cy5.5 anti‐CXCR5 (all from Miltenyi Biotec, Bergisch Gladbach, Germany). Cells were stained for 30 min at 4°C, measured with a FACSCanto II flow cytometer and analyzed using Kaluza Analysis version 2.1 (Beckman Coulter Life Sciences). Supporting information, Figure S3 reports the gating strategy for central and effector cell phenotyping panel, while Supporting information, Table S1 summarizes the main cell subsets identified from the analysis.
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