Fig 2: Gating strategy for identifying CXCR3 expression on monocytes using Kaluza software.a) First, we identified Singlets using forward height vs. area scatter. b) Next, we identified monocytes using a forward side scatter plot. c) Monocyte subsets were identified using the markers CD14 and CD16 (classical monocytes CD14+CD16-, intermediate monocytes CD14+CD16+, and non-classical monocytes CD14-CD16+). d) Finally, we used a negative isotype control (set to 1%) to distinguish the CXCR3 positive monocytes (shown in the overlay plot). The same procedure was done for lymphocytes, using the markers CD4+ and CD8+ for identifying CD4+ and CD8+ T cells.

Fig 3: Induction of Plasmablasts by a Live Attenuated Tetravalent DENV Vaccine(A) Plasmablasts were defined at the indicated times after vaccination by flow cytometry as CD38hiCD27hi cells within the CD19+CD20low/- B cell population in live CD3-CD14- PBMCs from TV003 vaccinees. A representative vaccine and placebo sample are shown. Values in the gate refer to the percentage of the CD19+CD20low/- parent population, and the blue-shaded values under the gate indicate the percentage of the plasmablasts of total CD19+ B cells.(B and C) Frequencies of plasmablasts (B) as a percentage of (C) total CD19+ B cells after vaccination with TV003 (n = 30 subjects). Boxplot areas show 25th-75th percentiles with the median as a thick line and whiskers indicating the 95% confidence interval and closed circles indicate each data point from each subject.(D) Mean plasmablast frequencies ± standard deviations (SDs) (red circles) and mean vaccine-associated viral titers ± SDs (blue squares) are plotted as a function of day after vaccination with TV003 (n = 20 subjects).(E and F) Correlation of day 21 plasmablast fold-rise (defined as plasmablast frequency at day 21 divided by the frequency at day 0) with (E) duration of viremia and (F) peak viral titer. Spearman R correlation coefficients and p values are reported. At least 2 technical replicates were performed for all of the viremia measurements.

Fig 4: Gating strategy for MFC-based assessment of minimal residual disease in autologous stem cell grafts. MCL cells were defined as single, viable cells negative for the lineage markers CD3/CD14/CD56 and the stem cell marker CD34, low/negative for CD23 and positive for CD5, CD19 and CD22. (A) Depiction of gating strategy in a control MCL sample (peripheral blood mononuclear cells). (B) The 1:100 000 dilution of control MCL sample in an autologous stem cell graft without measurable residual disease (UPN24). (C) Exemplary displays of positive MRD in autologous stem cell grafts of UPN31 (upper panels) and UPN10 (lower panels). (D) Exemplary displays of negative MRD in autologous stem cell grafts of UPN7 (upper panels) and UPN16 (lower panels). Abbreviations: FSC indicates forward scatter; lin; lineage; MFC, multiparameter flow cytometry; MRD, measurable residual disease; sLC, surface light chain; SSC, side scatter; UPN, uniform patient number.

Fig 5: Distinct phenotype and TCR repertoire for human iNKT cells from different anatomical locations.(A) Mean percentage of iNKT cells (CD1d-tet-PBS57+CD3+) from total CD3+B220-CD14- cells in blood or tonsils. (B–C) Flow-cytometry (left) and quantification (right) showing the percentage of CD4+, CD8+ and DN cells (B) or CD69+ and CD25+ cells (C) within the iNKT cell population in the depicted tissues. (D–E) Flow-cytometry (left) and quantification (right) showing the percentage of Vß11- iNKT cells within CD1d-tet-PBS57+Va24+ cells (D); or the percentage of Va24- cells within CD1d-tet-PBS57+Vß11+ cells (E). (A–E) Data are expressed as box-and-whisker diagrams depicting the median ± lower quartile, upper quartile, sample minimum and maximum. *p<0.05, ***p<0.001, two-tailed unpaired t-test. n = 10 samples per tissue from four independent experiments.Figure 7—source data 1.Distinct phenotype and TCR repertoire for human iNKT cells from different anatomical locations.