Apc anti cd45 clone hi30

Immediately after the 1 hour acclimation time to RT or 4°C, cells were placed in flow tubes at concentration of approximately 2×10⁶ cells per tube, incubated in fluorescently conjugated anti-human antibody cocktail for 20 minutes at ~4°C, washed in cold PBS, and fixed in cold 1% formaldehyde. Samples were analyzed on an LSR II flow cytometer (BD Biosciences). Single-color compensation and isotype controls were included for each experiment. Data analysis was performed using FlowJo 7.6.3 software (Tree Star). Gates were determined using fluorescence minus one controls. Percent of each population was used to calculate numbers/10⁶ CB cells. Human phenotyping markers included: FITC- human lineage cocktail 1 (CD3, CD14, CD19, CD20, CD56; catalog #340546; BD Biosciences), FITC or APC-anti-CD34 (clone 581; BD Biosciences), PE-anti-CD38 (clone HIT2; BD Biosciences), PE-CF594-anti-CD45RA (clone HI100; BD Biosciences), PE-Cy7-anti-CD90 (clone 5E10; BD Biosciences), PerCp-Cy5.5-anti-CD49f (clone WM53; BD Biosciences), APC-anti-CD45 (clone HI30; BD Biosciences) and APC-anti-CD184 (CXCR4; clone 12G5; BD Biosciences). HSCs were defined as lineage⁻ CD34⁺ CD38⁻ CD45RA⁻ CD90⁺ CD49f⁺, multipotent progenitors (MPPs) as lineage⁻ CD34⁺ CD38⁻ CD45RA⁻ CD90⁻ CD49f⁻, and progenitors as lineage⁻ CD34⁺ CD38⁺.^{13 (link)} See supplementary materials for mouse immunophenotyping.