Intracellular cytokine staining was performed as published previously [19 (link),24 (link)]. Briefly, splenocytes were stimulated with SARS-2 S peptide S268–276 (GYLQPRTFL) [19 (link)] or VACV peptide F226–34 (SPYAAGYDL) [27 (link)] to analyze SARS-2-S and MVA-specific CD8 T cells respectively. Non-treated cells and cells stimulated with PMA/ionomycin served as controls. Two hours after commencing the stimulation, Brefeldin A (Biolegend, San Diego, CA, USA) was added and cells were incubated for another 4 h. After stimulation, cells were stained extracellularly with anti-mouse CD3 phycoerythrin (PE)-Cy7, anti-mouse CD4 Brilliant Violet 421, anti-mouse CD8α Alexa Fluor 488 and purified CD16/CD32 (all from Biolegend, Supplementary Table S2 ). Cells were washed, fixed, permeabilized, and stained with anti-mouse IFN-γ allophycocyanin (APC) and anti-mouse TNF-α PE (both from Biolegend, Supplementary Table S2 ). Data were acquired using the MACSQuant VYB Flow Analyzer (Miltenyi Biotec, Bergisch Gladbach, Germany) or the NovoCyte Quanteon (Agilent Technologies, Waldbronn, Germany) and analyzed using FlowJo software (FlowJo LLC, Ashland, OR, USA).
Anti mouse cd8α alexa fluor 488
Manufactured by BioLegend
Sourced in United States
Anti-mouse CD8α Alexa Fluor 488 is a fluorochrome-conjugated antibody that binds to the CD8α protein on the surface of mouse T cells. It can be used for the identification and enumeration of CD8α-positive cells in flow cytometry analysis.
Automatically generated - may contain errors
Lab products found in correlation
Novocyte quanteon, by Agilent Technologies (1 mentions)
Cd16 cd32, by BioLegend (1 mentions)
Macsquant vyb flow analyzer, by Miltenyi Biotec (1 mentions)
Pe anti mouse tnf α, by BioLegend (1 mentions)
Brefeldin a, by BioLegend (1 mentions)
Software, by FlowJo (1 mentions)
Epcam apc, by BioLegend (1 mentions)
Apc cy7, by BioLegend (1 mentions)
Percp cy5, by BioLegend (1 mentions)
Anti mouse mhc 2 fitc, by Thermo Fisher Scientific (1 mentions)
Anti mouse cd45 brilliant violet 421, by BioLegend (1 mentions)
2 protocols using anti mouse cd8α alexa fluor 488
1
Intracellular Cytokine Staining for SARS-2 and MVA-Specific CD8 T Cells
2
Multiparameter Flow Cytometry of Immune Cell Subsets
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The following antibodies were used: anti-mouse CD3e-FITC, -PerCP-Cy5.5, or -APC-Cy7 (clone 145-2C11; BioLegend), CD4-APC or -PE-Cy7 (clone RM4-5; BioLegend), anti-mouse CD8α-Alexa Fluor 488 (clone 53-6.7; BioLegend), anti-mouse CD8β-PE (clone YTS156.7.7; BioLegend), anti-mouse CD27-Brilliant Violet 510 (clone LG.3A10; BioLegend), anti-mouse CD28-Brilliant Violet 421 (clone 37.51; BioLegend), anti-mouse CD45- Brilliant Violet 421 (clone 30-F11; BioLegend), anti-mouse CD274 (PD-L1)-PE (clone 10F.9G2; BioLegend), anti-mouse epithelial cell adhesion molecule (EpCAM)-APC (clone G8.8; BioLegend), anti-mouse MHC-II-FITC (clone M5/114.15.2; eBioscience), anti-mouse TCRβ- APC-Cy7 (clone H57-597; BioLegend) or anti-mouse γδTCR -PE-Cy7 (clone GL3; BioLegend). For scRNAseq analysis, anti-mouse Hashtags 1 to 4 (M1/42; 39-F11, Biolegend, 155861, 155863, 155865, 155867) were used.
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