Expression of galectin-9 in PBMCs is increased in early HIV-1 infection and correlates with HIV-1 viremia. Flow cytometry gating strategy to analyze intracellular expression of Gal-9 on (A) CD4+ and CD8+ T cells and (B) pDCs, monocytes or mDCs. Gates are set to exclude dead cells (viability marker+), and expression of Gal-9 on various lymphocyte subsets is defined by using appropriate fluorescence-minus-one controls. (C) Dot plots display the MFI of intracellular Gal-9 in the indicated immune cell subsets from 9 healthy (HIV-) and 5 untreated subjects with early infection as determined by flow cytometry. Horizontal lines indicate the median percentages. Differences where p < 0.05 are indicated. Histogram overlays display representative examples of intracellular Gal-9 expression in each lymphocyte subset analyzed in healthy controls (tinted) and in individuals with early HIV-1 infection (transparent) compared to the FMO (dotted line). PHI, early HIV-1 infection (D) Correlations between the logarithmic viral load and the MFI of intracellular Gal-9 in CD4 + T cells, CD8+ T cells, NK cells, monocytes, mDCS and pDCs from a subset of 32 HIV-1-positive individuals, including 4 subjects with early infection, 6 viremic controllers, 6 elite controllers, 9 subjects with chronic untreated, and 7 with chronic treated HIV-1 infection.