Donor lymphocyte infusion (DLI) into patients with a relapse of their leukemia or multiple myeloma after allogeneic stem cell transplantation (alloSCT) has been shown to be a successful treatment approach. The hematopoiesis-restricted minor histocompatibility antigens (mHAgs) HA-1 or HA-2 expressed on malignant cells of the recipient may serve as target antigens for alloreactive donor T cells. Recently we treated three mHAg HA-1- and/or HA-2-positive patients with a relapse of their disease after alloSCT with DLI from their mHAg HA-1- and/or HA-2-negative donors. Using HLA-A2/HA-1 and HA-2 peptide tetrameric complexes we showed the emergence of HA-1- and HA-2-specific CD8⁺ T cells in the blood of the recipients 5–7 weeks after DLI. The appearance of these tetramer-positive cells was followed immediately by a complete remission of the disease and restoration of 100% donor chimerism in each of the patients. Furthermore, cloned tetramer-positive T cells isolated during the clinical response specifically recognized HA-1 and HA-2 expressing malignant progenitor cells of the recipient and inhibited the growth of leukemic precursor cells in vitro. Thus, HA-1- and HA-2-specific cytotoxic T lymphocytes emerging in the blood of patients after DLI demonstrate graft-versus-leukemia or myeloma reactivity resulting in a durable remission. This finding implies that in vitro generated HA-1- and HA-2-specific cytotoxic T lymphocytes could be used as adoptive immunotherapy to treat hematological malignances relapsing after alloSCT.