Cd43 apc

To evaluate HE differentiation by flow cytometric analysis, hPSC-derived cells were dissociated by incubation in 0.1% collagenase for 2 h followed by TypLE Express for 8 min. Cells were mechanically dissociated and resuspended in 2% v/v FBS in Hank's buffered saline solution (HF). The following antibodies (obtained from BD Biosciences unless otherwise specified) were used at indicated dilutions: CD34-APC (1:100); CD144-PE (4:100); CD144-V450 (4:100); CD43-PE (3:100); CD45-PE-CY7 (4:100); CD4-PE (1:50) or CD34-PE (1:50); CD5-PE/Cy7 (1:200); CD7-AF700 (1:200); CD43-APC (1:200); CD45-APC/eF780 (1:200, eBiosciences); Ckit-APC (1:100); and PE-KDR (1:50). Control samples were prepared by incubating duplicate cell samples with respective fluorochome-labelled isotype antibodies. Samples were incubated with antibodies for 35 min on ice and then washed three times in cold HF. Viability discrimination was performed simultaneously using the viability stain 7-amino-actinomycin D at 1 μl ml⁻¹ (Molecular Probes). Samples were analysed on a Becton Dickinson FACS Fortessa machine using BD FACS Diva Software. Cells were sorted using a FACS Aria (BD Bioscience) cell sorter (Donnelly Centre for Cellular & Biomolecular Research).