Supplementary Figure 2: Factors that influence IgG3⁺IgM⁺ B cells.

(a) Longitudinal flow cytometry for IgG3 and IgM of CD20-gated B cells isolated from an HIV-infected individual during early viremia and one year later in chronic stage of viremia. Similarly, B cells from a chronically infected HIV-viremic individual were stained for IgG3 and IgM before and 10 years after sustained suppression of viremia by ART. (b) Comparison by stage of HIV infection and status of viremia of frequencies of IgG3⁺IgM⁺ and TLM B cells measured by flow cytometry, as well as of HIV viral load and CD4⁺ T-cell counts. Dotted line represents limit of detection. (c) Comparison by stage of HIV infection and status of viremia of mean fluorescence intensities (MFI) for IgG3 measured by flow cytometry of CD20-gated TLM and naïve B cells where IgG3-expressing B cells were excluded, as described in Methods and Supplementary Fig. 1c. (d) Comparison of IgG3 MFI on TLM versus naïve B cells of 92 HIV-viremic individuals, all stages of disease combined. Red horizontal bars represent medians; each symbol (b,c) represents an individual in early viremia (n = 16), chronic viremia (n = 73) or aviremia (n = 17); * P < 0.05, ** P < 0.01, *** P ≤ 0.001 and **** P < 0.0001; n.s., not significant (two-tailed Mann-Whitney test after obtaining significance by Kruskal-Wallis ANOVA test on full set (b,c) or two-tailed Wilcoxon matched-pairs signed rank test (d)). Data are representative of five (early to chronic viremic) and seven (chronic viremic to aviremic) individual experiments (a).