Bovine leukemia virus-infected cows with persistent lymphocytosis have an expanded population of B-lymphocytes in the peripheral blood that is sensitive to glucocorticoids in vitro and in vivo. We examined peripheral blood lymphocytes from cows with persistent lymphocytosis for the presence of specific glucocorticoid receptors. Steroid binding in intact cells was determined by a whole cell competitive binding assay using [3H]dexamethasone. The binding of the glucocorticoid to receptor was characterized in terms of affinity, specificity, and kinetics of the reaction. We found that peripheral blood lymphocytes from three cows with persistent lymphocytosis had 5000 to 6600 specific glucocorticoid-binding sites/cell. Compared with that reported for human lymphoid cells, glucocorticoid receptors in the bovine lymphocytes were found to have a greater affinity for the steroid with an association rate that was three times faster and a dissociation rate that was less than one-half of the former. We examined the biological half-life of hydrocortisone in the normal cow and found it to be 69.3 min, which is shorter than that reported for other domestic species and humans. The kinetics and affinity of the steroid binding may explain why in vivo glucocorticoid sensitivity was demonstrated in these animals despite the fact that elevated levels of plasma corticoids were not maintained. These results suggest that glucocorticoid sensitivity may be influenced by the nature of the binding reaction between steroid and receptor.