Figure 4. Mtb phagocytosis is anti-AM Ab mediated in TST⁺IGRA^– and TST⁺IGRA⁺ subjects but not those with TB.

(A) Effects of anti-AM IgG titers from 2 TST⁺IGRA⁺ (L) and 2 TST⁺IGRA^– (T) high-titer subjects before (+) and after (–) anti-AM Ab depletion. Columns represent mean of duplicates (circles). Notably, the depletion of the highest-titer serum (T2, a TST⁺IGRA^– subject) remained incomplete despite several depletion rounds. Sera were tested at 1:100 dilution. (B) IF microscopy of Mtb (H37Rv) with intact capsule showing reduced IgG binding with anti-AM Ab–depleted sera compared with nondepleted sera. The brightness and contrast of the images were adjusted with the same settings using ImageJ (NIH). (C) Decreases in THP-1 phagocytosis of Mtb opsonized with nondepleted versus anti-AM Ab–depleted heat-inactivated sera (10%). Columns and error bars represent mean and SD of triplicates (circles). (D and E) Reduction of Mtb phagocytosis in relation to anti-AM IgG titer depletion for TST⁺IGRA⁺ and TB sera. Sera were selected based on moderate to high anti-AM IgG titers (OD > 0.4 for TST⁺IGRA⁺ subjects, and OD > 0.5 for TB patients) and sufficient serum volume available to be able to assess the effects of reducing anti-AM Abs (TST⁺IGRA⁺ n = 10, TB n = 13). Spearman’s rank correlation. Duplicate or triplicate values are shown as circles. Results of assays repeated on separate days correlated highly and significantly (r > 0.8, P < 0.0001).