Anti h b m insulin apc conjugated rat igg2a

For identification of the insulin-positive population, 1 × 10⁶ iPSCs-derived IPCs were digested with trypsin, washed with PBS, and resuspended as single cells by incubation in Reagent 1: Fixation (Beckman Coulter) for 15 min. Then, the cells were washed once in PBS, incubated in Reagent 2: Permeabilization (Beckman Coulter) for 20 min, and washed once in PBS. Next, the cells were resuspended in PBS with primary antibody and incubated for 30 min. The cells were then washed with PBS twice and analyzed with the BD FACSCalibur system (BD Biosciences). The results were analyzed using FlowJo software. All procedures were carried out at room temperature. The primary antibody was anti-h/b/m insulin APC-conjugated rat IgG2A (R&D Systems). The isotype antibody was rat IgG2A control APC-conjugated.

For identification of the insulin positive cells, resuspended MDSCs-derived IPCs (1×10⁶ cells/mL) were incubated with the anti-h/b/m insulin APC-conjugated rat IgG2A (R&D Systems, MN, USA) for 30 min at room temperature in the dark. The insulin positive cells were detected using a flow cytometry (Beckman Coulter, Germany) and analyzed using CellQuest software (BD Biosciences, NJ, USA).