Journal: STAR Protocols

Article Title: CyTOF mass cytometry analysis of human memory CD4 + T cells and memory B cells

doi: 10.1016/j.xpro.2022.101269

Figure Lengend Snippet: Surface marker antibody cocktail

Article Snippet: 158 Gd-conjugated anti-human CD10 (clone HI10a) (1:50 dilution) , Fluidigm , Cat#3158011B.

Techniques: Marker

Journal: STAR Protocols

Article Title: CyTOF mass cytometry analysis of human memory CD4 + T cells and memory B cells

doi: 10.1016/j.xpro.2022.101269

Figure Lengend Snippet: CyTOF data analysis (A) PBMCs from four healthy donors were stained with a panel of metal-labeled antibodies and analyzed by CyTOF. After data pre-processing, manual gating was used to identify classical MBCs (CD3 - CD19 + CD20 + CD10 - CD27 + CD21 + ), atypical MBCs (CD3 - CD19 + CD20 + CD10 - CD27 - CD21 - ) and activated MBCs (CD3 - CD19 + CD20 + CD10 - CD27 + CD21 - ). tSNE analysis was then performed in Cytobank ( Kotecha et al., 2010 ) before self-organizing maps (SOMs) were generated using hierarchical consensus clustering on the tSNE axes. (B) tSNE and FlowSOM analysis of classical MBCs. (C) tSNE and FlowSOM analysis of atypical MBCs. (D) tSNE and FlowSOM analysis of activated MBCs. (E) A similar approach was used to analyze the memory CD4 + T cell compartment. Manual gating was used to identify T H1 memory CD4 + T cells (CD19 - CD3 + CD4 + CD45RA - CCR6 - CXCR3 + ), T H2 memory CD4 + T cells (CD19 - CD3 + CD4 + CD45RA - CCR6 - CXCR3 - ), T H17 memory CD4 + T cells (CD19 - CD3 + CD4 + CD45RA - CCR6 + CXCR3 - ) and circulating memory T FH cells (CD19 - CD3 + CD4 + CD45RA - CXCR5 + ) before tSNE analysis and FlowSOM clustering was performed for each memory CD4 + T cell sub-set. (F) tSNE and FlowSOM analysis of T H1 memory CD4 + T cells. (G) tSNE and FlowSOM analysis of T H2 memory CD4 + T cells. (H) tSNE and FlowSOM analysis of T H17 memory CD4 + T cells. (I) tSNE and FlowSOM analysis of circulating memory T FH cells. The tSNE plots in the upper left panel display cell density and represent the pooled data from four healthy donors, while the upper right panel shows a projection of the FlowSOM clusters on the tSNE plot. Heatmaps shows the mean marker expression for each FlowSOM cluster.

Article Snippet: 158 Gd-conjugated anti-human CD10 (clone HI10a) (1:50 dilution) , Fluidigm , Cat#3158011B.

Techniques: Staining, Labeling, Generated, Marker, Expressing

Journal: STAR Protocols

Article Title: CyTOF mass cytometry analysis of human memory CD4 + T cells and memory B cells

doi: 10.1016/j.xpro.2022.101269

Figure Lengend Snippet:

Article Snippet: 158 Gd-conjugated anti-human CD10 (clone HI10a) (1:50 dilution) , Fluidigm , Cat#3158011B.

Techniques: Recombinant, Saline, Staining, Software, Cytometry