An understanding of the immune response to rotavirus is needed to develop effective prophylaxis. There is evidence that cell-mediated responses may be involved and to extend these observations, rotavirus antigen and the three major T cell subsets, BoCD4 +, BoCD8 +, and BoWC1 + γ/ δ lymphocytes were immunostained in tissue sections from calves killed at 2, 4, 6, 8 and 10 days post inoculation and quantified by image analysis. It was established that in control calves, BoCD4 + lymphocytes were predominantly in the lamina propria, while the majority of BoCD8 + and BoWC1 + γ/ δ lymphocytes were in the epithelium. Rotavirus infection was seen throughout the small intestine with the greatest amount of viral antigen detected at 4 days post inoculation in the mid and distal small intestine. Increased numbers of all subsets were detected; small increases in intraepithelial BoCD4 + and BoWC1 + γ/ δ T lymphocytes were observed especially in the distal small intestine, while larger increases in BoCD8 + cells were detected in the epithelium and lamina propria of the proximal, mid and distal small intestine. The timing and location of these increases in T lymphocyte subsets is indicative of a specific immune response involving BoCD8 + and BoWC1 + γ/ δ T lymphocytes.