Cell surface molecules aberrantly expressed or overexpressed by myeloid leukemic cells represent potential disease-specific therapeutic targets for antibodies. MUC1 is a polymor-phic glycoprotein, the cleavage of which yields two unequal chains: a large extracellular a subunit containing a tandem repeat array bound in a strong noncovalent interaction to a smaller b subunit containing the transmembrane and cytoplasmic domains. Because the a-chain can be released from the cell-bound domains of MUC1, agents directed against the a-chain will not effectively target MUC1 + cells. The MUC1 SEA (a highly conserved protein module so called from its initial identification in a sea urchin sperm protein, in enterokinase, and in agrin) domain formed by the binding of the a and b chains represents a stable structure fixed to the cell surface at all times. DMB-5F3, a partially human-ized murine anti-MUC1 SEA domain monoclonal antibody, was used to examine MUC1 expression in acute myeloid leukemia (AML) and was found to bind acute myelomonocytic and monocytic leukemia (AML-M4 and AML-M5) cell lines. We also examined monocytic neoplasms freshly obtained from patients including chronic myelomonocytic leukemia and juvenile myelomonocytic leukemia, which were found to uniformly express MUC1. CD34 + / lin − /CD38 − or CD38 + presumed leukemic stem cell populations from CD34 + AML and CD34 − CD38 − or CD38 + populations from CD34 − AML were also found to express MUC1, although at low percentages. Based on these studies, we generated an anti-MUC1 immuno-toxin to directly gauge the cytotoxic efficacy of targeting AML-bound MUC1. Using single-chain DMB-5F3 fused to recombinant gelonin toxin, the degree of AML cytotoxicity was found to correlate with MUC1 expression. Our data support the use of an anti−MUC1 SEA module−drug conjugates to selectively target and inhibit MUC1-expressing myelomo-nocytic leukemic cells.