Stronger expansion of OT-II CD4⁺ T cells in mice immunized with OVA plus CpG than with OVA plus CL097.

Mice were adoptively transferred with 1 x 10⁶ splenocytes obtained from OT-II mice and, 24 h later, immunized with OVA protein (25 μg) with or without CpG (20 μg) or CL097 (20 μg) formulated in IFA. Mice were sacrificed 11 days post immunization, and the number of expanded OT-II CD4⁺ T cells were quantified by cell surface staining of CD4 and CD45.1 followed by flow cytometric analysis. (A) A series of gating steps depicted in arrows involved gating on total leukocytes (FSC-A/SSC-A), on singlets (FSC-A/FSC-H), and on live cells (7-AAD/SSC-A). (B) Each FACS plot is an analysis of live splenocytes for one of eight mice within a group. Cells double positive for CD4 and CD45.1 demarcated in the plot are OT-II CD4⁺ T cells. Numbers are the percentages of the OT-II CD4⁺ T cells in spleens. (C) The total numbers of OT-II CD4⁺ T cells in spleens were calculated based on the percentages of OT-II CD4⁺ T cells and known numbers of splenocytes. Statistical differences were analyzed by one-way ANOVA between treatment groups. Data shown are representative of three independent experiments. *p<0.002.