Terminal deoxynucleotidyl transferase (TdT) is a useful marker for normal lymphocyte precursors and acute lymphoblastic leukemia (ALL). Our previous studies, however, have shown that for monitoring minimal residual disease in the circulation, assay for TdT alone is not sufficiently specific to distinguish leukemia cells from the background of rare normal blood TdT+ cells. In an attempt to increase specificity for leukemic cells, we have used double and triple immunophenotypic analysis to characterize normal circulating and bone marrow TdT+ cells. Overall, normal TdT+ cells were about 1000-fold more frequent in the marrow than in the blood. More than 75% of TdT+ cells in both the blood and marrow expressed the CD34, CD22, and HLA-DR antigens. However, circulating TdT+ cells infrequently expressed CD19 (4.5%) and CD9 (2.3%), compared with their marrow counterparts (74% and 47%, respectively). The brightly staining CD10+ phenotype, frequently associated with ALL blasts, was significantly less common among normal blood (5.7%) than marrow (31%) TdT+ cells. Although T-lineage markers were rarely expressed on TdT+ cells in either site, CD7+ cells were far more prevalent within the circulating TdT+ subset (4%) than among the marrow population (less than 0.2%). The results suggest a selective release of lineage-uncommitted and/or thymus-destined TdT+ cells from the marrow into the circulation. Moreover, since CD19, CD9, and high-density CD10 are frequently found on ALL blasts, staining for these markers on TdT+ cells in the circulation should improve the specificity of assay for residual common ALL cells. Likewise, assay for CD5+ and possibly CD7+ TdT+ cells in either marrow or blood should provide a very sensitive method of detection of T-ALL blasts.