Classical immunotherapeutic approaches to glioblastoma multiforme (GBM) have shown mixed results, and therapies focused on innate lymphocyte activity against GBM have not been rigorously evaluated. We examined peripheral blood lymphocyte phenotype, γδ T-cell number, mitogenic response, and cytotoxicity against GBM cell lines and primary tumor explants from GBM patients at selected time points prior to and during GBM therapy. Healthy volunteers served as controls and were grouped by age. T-cell infiltration of tumors from these patients was assessed by staining for CD3 and T-cell receptor γδ. Our findings revealed no differences in counts of mean absolute T-cells, T-cell subsets CD3+CD4+ and CD3 +CD8+, and natural killer cells from healthy volunteers and patients prior to and immediately after GBM resection. In contrast, γδ T-cell counts and mitogen-stimulated proliferative response of γδ T-cells were markedly decreased prior to GBM resection and throughout therapy. Expanded/activated γδ T-cells from both patients and healthy volunteers kill GBM cell lines D54, U373, and U251, as well as primary GBM, without cytotoxicity to primary astrocyte cultures. Perivascular T-cell accumulation was noted in paraffin sections, but no organized T-cell invasion of the tumor parenchyma was seen. Taken together, these data suggest that γδ T-cell depletion and impaired function occur prior to or concurrent with the growth of the tumor. The significant cytotoxicity of expanded/activated γδ T-cells from both healthy controls and selected patients against primary GBM explants may open a previously unexplored approach to cellular immunotherapy of GBM. Copyright 2009 by the Society for Neuro-Oncology.