Hematopoietic reconstitution by bone marrow transplantation (BMT) is used as therapy for the treatment of various malignancies and genetic blood disorders. Allogeneic BMT is the most common application of this treatment but is frequently associated with graft-versus-host disease (GVHD). Recent clinical studies have shown that sibling transplant using umbilical cord blood (UCB) is an acceptable alternative to BMT and may involve fewer problems with GVHD. We have investigated the in vitro alloreactive capacity of UCB as it relates to allogeneic transplantation. Initial screening assays demonstrated that UCB T cells were functionally immature. It was not possible to generate significant levels of alloantigen-specific cytotoxic T lymphocytes (CTL) in either primary or secondary mixed lymphocyte cultures. Limiting dilution analyses revealed that cord blood T cells were 10-1000 x less alloreactive in terms of proliferative T cells (PTLp) and cytotoxic T cells (CTLp) compared with adult peripheral blood lymphocytes (PBL). However, UCB was equivalent to adult PBL in terms of natural killer (NK) and lymphokine-activated killer (LAK) cell precursors. Analysis of cells from alloantigen-stimulated MLC revealed that UCB generated primarily CD4+ and CD16+ cells that made little or no IL-4, IL-6, TNF-alpha or IFN-gamma on antigenic stimulation. Cold target inhibition analyses revealed that alloantigen-stimulated cord blood T cells had a fine specificity similar to NK cells. From these in vitro results cord blood would seem to be unlikely to mediate severe GVHD reactions in vivo and should be suitable for allogeneic transplantation.