Adoptive transfer of virus-specific T cells offers the potential for accelerating reconstitution of antigen-specific immunity and limiting the morbidity and mortality of viral infections following allogeneic haematopoietic stem cell transplantation. However, the logistics of producing virus-specific T cells and the risk of inducing graft-versus-host disease secondary to the infusion of alloreactive clones have limited the application of cellular therapies. We report the results in patients of pre-emptive and prophylactic therapy with cytomegalovirus-specific T cells. Cells were administered at early time points following transplantation (when the risk of GVHD is greatest) either prophylactically or following the detection of CMV DNA by a PCR-based surveillance technique. Massive in vivo expansions of CMV-specific cytotoxic T-lymphocytes (3-5 log) were observed in patients within days of adoptive transfer. Viral titers were decreasing within 5 days, in some patients the T-cell receptor CDR3 lengths of CMV-specific CTL expanding in vivo were identical to those of the transferred cells. A low incidence of late cytomegalovirus reactivation was seen and no significant toxicities were observed. Our findings indicate that application of cell lines generated by either short-term in vitro cultures or by direct selection using gamma-capture, which allow expansion of both CD4(+) and CD8(+) virus-specific T cells, is both feasible and effective in a clinical environment. These simple in vitro methodologies should allow widespread application of adoptive transfer of virus-specific T cells.