The thymus is critical for the establishment of a functional and self-tolerant adaptive immune system but involutes with age, resulting in reduced naive Tcell output. Generation of a functional human thymus from human pluripotent stem cells (hPSCs) is an attractive regenerative strategy. Direct differentiation of thymic epithelial progenitors (TEPs) from hPSCs has been demonstrated invitro, but functional thymic epithelial cells (TECs) only form months after transplantation of TEPs invivo. We show the generation of TECs invitro in isogenic stem cell-derived thymic organoids (sTOs) consisting of TEPs, hematopoietic progenitor cells, and mesenchymal cells, differentiated from the same hPSC line. sTOs support Tcell development, express key markers of negative selection, including the autoimmune regulator (AIRE) protein, and facilitate regulatory Tcell development. sTOs provide the basis for functional patient-specific thymic organoid models, allowing for the study of human thymus function, Tcell development, and transplant immunity.