Macs bead sorting system

Cells (2.5 × 10⁶ cells/ml) were stained with antibodies against TYR (Abcam), TYRP1 (Sigma), c-Kit (eBioscience) and PDGFRA (BioLegend). To detect intracellular proteins, staining was carried out on cells fixed with 4% paraformaldehyde (Electron Microscopy Sciences) in PBS. Staining was performed in PBS with 2% FBS. Stained cells and GFP⁺ cells were analyzed using an LSRII flow cytometer (BD). For FACS sorting, cells were sorted at a concentration of 10⁶ cells/ml in PBS/2% FBS using a FACSAriaTMII (BD) cell sorter (Upenn Flow Cytometry Facility). PDGFRA⁺/c-Kit⁻ fibroblasts were sorted from P1 fetal hFs and cultured as P0. For magnetic bead sorting, the Miltenyi MACS bead sorting system was used according to the manufacturer’s guidelines. Data were analyzed using FlowJo software (Treestar). The working solutions of antibodies are shown in Supplementary Table S3.

Cells were stained at a concentration of 2.5 × 10⁶ cells/ml with antibodies against CD200, ITGA6, SSEA3, KRT14 and KRT15. For cell-surface markers, staining was carried out in PBS with 2% FBS. For intracellular proteins, staining was carried out on cells fixed with 4% paraformaldehyde (Electron Microscopy Sciences) in PBS. Staining was done in PBS with 2% FBS. Stained cells were analyzed using an LSRII flow cytometer (BD Biosciences). For Fluorescence Activated Cell Sorting, the cells were sorted at a concentration of 10⁶ cells/ml in PBS/2% FBS using a FACS AriaTMII cell sorter (BD Biosciences). For magnetic bead sorting, the Miltenyi MACS bead sorting system was used according to the manufacturer's guidelines and sorting conditions. Data were analyzed using FlowJo software (Treestar). A comprehensive list of antibodies is described in Supplementary Table 2.