The EphA2 receptor tyrosine kinase is selectively expressed on the surface of many different human tumors. We have previously shown that tumor cells can be targeted by EphA2 monoclonal antibodies and that these antibodies function, in part, by inducing EphA2 internalization and degradation. In this report, we describe the isolation and characterization of a fully human monoclonal antibody (1C1) that selectively binds both the human and rodent EphA2 receptor. After cell binding, the antibody induces rapid tyrosine phosphorylation, internalization, and degradation of the EphA2 receptor. Because monoclonal antibodies that selectively bind tumor cells and internalize provide a vehicle for targeted delivery of cytotoxics, 1C1 was conjugated to the microtubule inhibitor monomethylauristatin phenylalanine using a stable maleimidocaproyl linker. The anti-EphA2 antibody-drug conjugate [1C1-maleimidocaproyl-MMAF (mcMMAF)] stimulated the activation of caspase-3/caspase-7 and the death of EphA2-expressing cells with IC50 values as low as 3 ng/mL. Similarly, the conjugate induced degradation of the EphA2 receptor and inhibited tumor growth in vivo. Administration of 1C1-mcMMAF at doses as low as 1 mg/kg once weekly resulted in significant growth inhibition of EphA2-expressing tumors without any observable adverse effects in mouse xenograft and rat syngeneic tumor models. Our data support the use of an antibody-drug conjugate approach to selectively target and inhibit the growth of EphA2-expressing tumors.