CD34+ HSPCs can be isolated from either bone marrow, thymus, fetal liver, umbilical cord blood, peripheral blood or in vitro differentiated from ESCs and iPSCs. These HSPCs can be expanded and manipulated ex vivo before subsequent application in in vitro T cell differentiation models. Green and purple sticks represent DLL1/4 Notch ligand and VCAM-1, respectively. The OP9 stromal cells and microbeads are depicted as brown cells and silver spheres, respectively. ESC: embryonic stem cell; iPSC: induced pluripotent stem cell; HSPC: hematopoietic stem and precursor cell; FTOC: fetal thymic organ culture; RTOC: reaggregated thymic organ culture; TEC: thymic epithelial cell; OP9-DLL1/4: OP9-Delta-like ligand 1/4; ATO: artificial thymic organoid; SP8: CD8 single positive; VCAM-1: vascular cell adhesion molecule 1; UCB: umbilical cord blood; mPBL: mobilized peripheral blood; PSC: pluripotent stem cell. (Created with BioRender.com).