Mlxeq four calibration beads

Antibodies used for mass cytometry are shown in Table 2. Directly conjugated antibodies were purchased from Fluidigm and purified antibodies were ordered from the companies listed in Table 2. Conjugations were performed with the Maxpar X8 Multi-Metal Labeling Kit (Fluidigm) according to the manufacturer’s instructions. Samples were stained for CyTOF mass cytometry, as described previously^{25 (link), 26 (link)}, and stained with the antibody cocktail shown in Table 2. Prior to surface staining, one low CVD sample and one high CVD sample, in addition to a staining healthy control, were barcoded with CD45 (low CVD-CD45–89Y; High CVD-CD45–147Sm; Healthy-CD45–115Ln). Cells were resuspended at 2×10⁶ cells/mL in 0.1mLxEQ^™ Four Calibration Beads (Fluidigm). Batch effects and signal drifting were minimized by tuning every 6hrs during long runs. Data was normalized using the Matlab-based NormalizerR2013a_Win64.

Conjugated antibodies were purchased from Fluidigm and purified antibodies were ordered from companies as listed in Table IV. Metal conjugations were performed as needed with the Maxpar X8 Multi-Metal Labeling Kit. Samples were stained for CyTOF mass cytometry according to our previously published methods.^{25 (link)} Low CVD (CD45–89Y), high CVD (CD45–147Sm), and staining control (CD45–115Ln) PBMCs were barcoded (S.Fig. 3). Age-matched healthy and moderate CVD samples were separately stained by mass cytometry and barcoded with CD45–89Y and CD45–147Sm, with CD45–115Ln as a staining control. Cells were resuspended at 2×10⁶ cells/mL in 0.1mLxEQ™ Four Calibration Beads (Fluidigm). Tuning was performed every 6hrs. Data was normalized using the Matlab-based NormalizerR2013a_Win64.