Abstract Glucocorticoid receptor levels were quantitated in leukemic blasts from bone marrow aspirates of 174 children with acute lymphocytic leukemia. Using a whole-cell assay, we found that [3H]dexamethasone binding had the following characteristics: (a) reached a steady state in 30 min at 22° and was stable for at least 2 hr; (b) was linear with cell number between 1 and 6 × 106 cells/assay; (c) was higher in freshly prepared blasts compared to blasts stored for 18 hr at 4 or 22°; (d) was specific for glucocorticoids; (e) had a Kd of ≈1 × 10-8m; and (f) required a postincubation washing step to maximize sensitivity and decrease nonspecific binding. Glucocorticoid receptor levels in patient samples exhibited a wide range from 2,248 to 79,364 sites/cell (median, 18,123). A lower level was correlated with the following biological and clinical characteristics at diagnosis: high leukocyte count; positive sheep erythrocyte rosette test; T-cell surface antigens; presence of mediastinal mass; age 10 years; central nervous system leukemia; and black race. There was no correlation of receptor levels with in vitro [3H]thymidine labeling indices, terminal deoxynucleotide transferase levels, FAB classification, or sex. For 43 patients treated on Total Therapy Study IX (follow-up duration from 24 to 33 months), a lower receptor level ( 100 × 103/cu mm, positive erythrocyte rosette test, central nervous system involvement, and mediastinal mass) were excluded, lower glucocorticoid receptor levels were still associated with higher risk of relapse (p = 0.007). We conclude that glucocorticoid receptor levels may be an important prognostic indicator in childbood acute lymphocytic leukemia.