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Additional Supporting Information may be found in the online version of this article.

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CYTO_22071_sm_SuppFig1.tif6930KSupplemental Figure 1. Reagent titrations . Reagents were titrated on 4 ml whole peripheral blood, RBC-lysed, from healthy donors. Single FCS files were concatenated to show titrations for each reagent in a single plot. Antibodies and reagents concentrations are displayed along the x-Axis. CD146 and CD106 titrations were carried out by adding CD45 and CD34 antibodies and analyzing CD45-/CD34+ events. Concentrations used in OMIP are highlighted in red.
CYTO_22071_sm_SuppFig2.tif3177KSupplemental Figure 2. Positive controls. Reagent titrations were performed on other cell types. A CD34-ECD staining (20/100) on cord blood cells. B Staining of CD106 PE-Cy5 (2/100) on TNF-alpha-activated (16 H, 1 ng / ml) HUVEC cells (left plot) compared to control (CTRL – right plot). C Internal controls for CD34 and CD117: CD45+/CD34+/CD117+ cells represent the well-known circulating haemopoietic stem cell compartment. D Internal control for CD146 staining: CD45+/CD146+ cells are peripheral activated-T lymphocytes.
CYTO_22071_sm_SuppFig3.tif11408KSupplemental Figure 3. Gating strategy and FMO controls. A Cells were not gated on morphological parameters, only artifacts in the upper left and lower right corner were excluded. Aggregates were gated out on a FSC-Area (FSC-A) vs. FSC-High (FSC-H) dot plot. Debris, dead cells, microparticles, and platelets were excluded by gating Syto16+ cells and NiRed- cells. Leucocytes were excluded by gating CD45-, CD45dim and CD31+ cells. CD34+, CD146+, CD106+ and CD117+ cells were separately gated. B Appropriate FMO controls are shown. Dot plots show 8,000 events each, whereas pseudocolour plots and low resolution zebra plots show all analyzed events.
CYTO_22071_sm_SuppFig4.tif4919KSupplemental Figure 4. Spillover into all detectors. CompBeads were labeled individually with each of 6 fluorochromes and blood samples were stained with Syto16 or NiRed. The percentage of light from the excitation of the primary fluorochrome contaminating each of the other detectors was calculated, as suggested by Perfetto SP et al. (Nat Rev Immunol, 2004). Each bar represents the percentage of the signal subtracted in the given detector, following the compensation procedure.
CYTO_22071_sm_SuppTab1.tif3821KSupplemental Table 1. Instrument configuration. The panel was optimized for a FACSCanto II with the listed optical elements. Long pass dichroic filters were used throughout the instrument. Light is filtered to each photomultiplier tube such that only wavelengths lower than long pass filter and within the band pass are detected. DPSS – Diode Pumped Solid State, HeNe – Helium-Neon, Cy – cyanine, ECD – energy-coupling dye, LP – long pass, NiRed – LIVE/DEAD fixable near-infrared dead cells stain, PE – R-phycoerythrin.
CYTO_22071_sm_SuppTab2.tif6501KSupplemental Table 2. Commercially available reagents used in OMIP-011.
CYTO_22071_sm_SuppTab3.tif8048KSupplemental Table 3. Reagents tested, but not used. FITC – fluorescein isothiocyanate, H7 - Hilite®7, PerCP - peridinin chlorophyl protein-conjugated.
CYTO_22071_sm_SuppInfo.doc36KSupporting Information

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