EGFRvIII is the most common deletion variant of the epidermal growth factor receptor and is found in cancers of the brain, breast, ovary, and lung. The complete absence of the receptor in healthy tissues makes it an ideal tumor marker. We sought to design a peptide ligand against EGFRvIII for development as a diagnostic imaging agent. We used the concept of hydropathic complementarity to search for sequences whose amino acid sidechains display a reciprocal pattern of hydropathicity to those of the deletion junction of EGFRvIII. The resulting peptide (PEPHC1) was synthesized and tested for binding to EGFRvIII and EGFR. In in vitro assays, PEPHC1 bound the recombinant EGFRvIII extracellular domain or full-length EGFRvIII solubilized from cell membranes in preference to native EGFR. These results demonstrate the utility of hydropathic complementarity as a basis for the design of highly specific ligands that may prove useful as tumor-targeting agents.