Fitc conjugated cd38

Evaluation of Breg cells frequencies by flow cytometry, circulating Breg cells were detected using FITC-conjugated-CD38, PE-conjugated-CD24 (BD Bioscience, USA), and PerCP-conjugated CD19 (BD Bioscience, USA).100 μl of blood sample was incubated with 10 μl of CD24, CD38, and CD19 for 20 minutes at 4 C in the dark. Following incubation, red blood cell lysis, washing, and analysis by FACS Calibur flow cytometry with Cell Quest software (Becton Dickinson Biosciences, USA) was done. An isotype-matched negative control was used for each sample. A Forward and side scatter histogram was used to define the lymphocytes. Then CD19+ B cells were gated. Then the expression of CD38 and CD24 on the CD19+B cells was revealed. Regulatory B cells were specified as CD19⁺CD24^highCD38^high cells (Figure 1).

The iHPCs were dissociated into single cells with TrypLE™ Express and counted, and 5 × 10⁴ cells were suspended in 100 μL DPBS with 5% BSA. Then the cells were incubated with BV421-conjugated anti-human CD34, FITC-conjugated CD38, BV510-conjugated CD43, and APC-conjugated CD41a (BD Biosciences, Franklin Lakes, NJ, USA) at 4 °C for 30 min in the dark. The stained cells were washed twice using DPBS and characterized by flow cytometry. The iMKs were incubated with BV421-conjugated anti-human CD34, BV510-conjugated CD43, and APC-conjugated CD41a (BD Biosciences, Franklin Lakes, NJ, USA) at 4 °C for 30 min in the dark and analyzed by flow cytometry.