Anti human cd45 pe

First, approximately 1 × 10⁵ cells were collected from the culture, centrifuged for 5 min at 300 g to remove the supernatant, and washed twice with 2 ml 1 × Dulbecco’s phosphate-buffered saline (DPBS; catalog No. 14190136, Gibco) solution containing 2% fetal bovine serum (Catalog No. 16000044, Gibco) and 2 mM EDTA. After centrifugation, the cells were resuspended in 50 μl of 1 × DPBS buffer, and the antibodies were added, followed by incubation at 4 °C for 10 min in the dark. The cells were washed twice by adding 2 ml of 1 × DPBS buffer after incubation and then resuspended in 200 μl of 1 × DPBS buffer to prepare a cell suspension for the assay. The BD FACSAria II instrument was used for flow cytometry analysis, and the data analysis was performed with Flowjo software (v.7.6, Three Star). The antibodies used in this study included anti-human CD34-FITC (Catalog No. 130-098-142, Miltenyi Biotec, Bergisch Gladbach, Germany), anti-human CD45-PE (Catalog No. 170-081-061, Miltenyi Biotec), anti-human CD71-PE (Catalog No. 130-099-219, Miltenyi Biotec), and anti-human CD235a-APC (Catalog No. 130-118-493, Miltenyi Biotec).