We focused on inflammatory and immune activation markers that have been associated with stroke and cardiovascular disease in HIV and non-HIV populations[28 (link)–37 (link)]. We tested inflammatory and immune activation markers in cryopreserved biospecimens from the most recent WIHS visit preceding the TCD study. IL-6 and CRP were measured in plasma samples using a multiplex electrochemiluminescence assay (Meso Scale Discovery, MD, USA). Soluble CD14 (R&D Systems, MN, USA) and CD163 (Aviscera Bioscience, CA, USA) were measured by ELISA. Details of peripheral blood mononuclear cell (PBMC) laboratory testing have been described previously[38 (link)]. Cryopreserved PBMCs were thawed in batches and stained with viability dye LIVE/DEAD® Fixable Blue Dead Cell Stain Kit (Life Technologies, NY, USA). Cells were washed and stained with fluorescent conjugated antibodies for cell surface markers. To measure CD4+ and CD8+ T cell activation and identify monocytes, PBMCs were stained as described previously[38 (link)]. Subpopulations of monocytes were evaluated with stains for anti-CD14 FITC (eBioscience, CA, USA), anti-CD16 PE-Cy7 (Biolegend, CA, USA), anti-CCR2 PerCPCy5.5 (Biolegend), anti-CX3CR1 APC (Biolegend), anti-CD163 PE (R&D systems) and anti-CCR5 APC-Cy7 (BD, NJ, USA). Cellular markers were detected by flow cytometry using LSRII flow cytometer (BD).