Fig. 3. Anti-PstS1 mAbs inhibit Mtb in culture.

a Upper panel: gating strategy of H37Ra-infected macrophages, mCherry positive. PMA-differentiated THP-1 cells preincubated with mAbs p4-163, p4-170, p4-36, p4-141, and the isotype control mAb mGO53²⁰. For each treatment n = 90,000 cells were analyzed by flow cytometry. Lower panel: histograms showing the frequencies of intracellular antibody-bound bacteria as detected by anti-human VioBlue antibody staining. For each mAb, the binding histogram is shown in red and compared to the isotype control histogram, which is depicted in a gray overlay. b, c Activity of anti-PstS1 mAbs at indicated concentrations in a human whole blood mycobacterial growth inhibition assay (MGIA) after 96 h of infection with BCG or pathogenic Mtb, respectively. CFU was determined in n = 3 biological repetitions. d Activity of anti-PstS1 mAbs (5 µg/ml) used as IgG1 (named “WT”, full columns) or as N279A Fc variants (named “NA”, empty columns) in MGIA. Black and clear shapes correspond to two independent experiments. CFU was determined in n = 5–6 biological repetitions. e Activity of anti-PstS1 mAbs (5 µg/ml) in MGIA following depletion of CD3+ T cells, CD4+ T cells, CD8+ T cells, blockade of MHC II (anti-HLA), and CD16 or CD32 or both (anti-CD16, anti-CD32, marked with a red rectangle). Black, cayenne, and clear shapes correspond to three representative independent experiments. In each experiment, the data points were compared to average CFU infection in PBS, which was normalized to 1. CFU was determined in n = 4–10 biological repetitions. All error bars are represented as mean ± SD. Significance was determined using a one-tailed unpaired t test (b), two-tailed unpaired t test (c, d) for black shapes (d), or one-way ANOVA with Tukey’s multiple-comparison test (e). All statistical analyses are relative to PBS. ns no significance. Data are representative of at least two independent experiments.