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. 2021 May 6;11:9686. doi: 10.1038/s41598-021-88956-4

Figure 1.

Figure 1

IgE-mediated mast cell activation does not result in increased numbers of endogenous antigen-specific CD4+ T cells. (A) Strategy for intradermal ear immunization to enumerate 2W1S-specific CD4+ T cells upon IgE/OVA-induced mast cell activation. (B) Gating strategy and representative flow plots of antigen-specific CD4 + T cells in the cervical draining lymph nodes (CLNs) 7 days post immunization. 2W1S-specific CD4 + T cells post enrichment were identified as lymphocytes by forward and side scatter properties and then doublets were excluded based on side scatter width (SSC-W). Singlets were stained for T cells by eliminating T cell lineage negative cells (CD11b-, CD11c-, F4/80-, CD19-), and gating on CD3ε + cell. Helper T cells were gated as CD4 + CD8α- cells. Finally, antigen experienced 2W1S specific cells were gated as CD44hi and I-Ab :2W1S-APC + . Percentages in each gate are indicated. (C) Representative flow plot of 2W1S-specific CD4+ T cells in the cervical draining lymph nodes (CLNs) 7 days post immunization for each group is shown. Percentage of 2W1S-specific CD4+ T cells post tetramer enrichment was indicated. (C) The total number of 2W1S-specific CD4+ T cells in the CLNs 7 days post immunization. Results are shown as mean ± SEM from 4 independent experiments n = 11–12 mice per group. Statistical analysis was performed using a Kruskal–Wallis test with Dunn’s correction.