The thymus is the central hub of T lymphocyte differentiation, where epithelial cells guide the process of their maturation. Progenitors and stem cells are found in niches deep within the tissues of an adult organism. Their function is tissue regeneration and maintenance of homeostasis. The existence of stromal stem cells within the postnatal mouse thymus has been shown, but these have not been proven to exist in the human thymus. The aim of this research was to find epithelial stem cells within the human thymus. Sequential tissue digestion and plating the obtained cells in a manner suited for 3D cultures, we established a method that consistently leads to a formation of organoids indicating the presence of stem cells. The organoids are not mere cell aggregates, they are formed from individual cells with the capability of self-renewal, as well as other properties of stem cells, such as maintaining their immature state while giving rise to differentiated cells. The removal of lymphocytes from the cell suspension by either magnetic depletion or incubation with 2'-deoxyguanosine leads to an increase of organoid number. The cells within the organoid express genes necessary for thymic epithelial function, but also genes active during thymic organogenesis. The expression of protein markers of mature epithelial cells has been found on a smaller proportion of organoid cells. The organoids are capable to lead the differentiation of a hematopoietic stem cell into mature single positive T lymphocytes.