Peptide-modified cryptophane enables sensitive detection ofproteinanalytes using hyperpolarized ¹²⁹Xe NMR spectroscopy. Here we report improved targeting and delivery of cryptophane to cells expressing α_vβ₃ integrin receptor, which is overexpressed in many human cancers. Cryptophane was functionalized with cyclic RGDyKpeptide and Alexa Fluor 488dye, and cellular internalization was monitored by confocal laser scanning microscopy. Competitive blocking assays confirmed cryptophaneendocytosis through an α_vβ₃ integrin receptor-mediated pathway. The peptide–cryptophane conjugate was determined to be nontoxic in normal human lung fibroblasts by MTT assay at the micromolar cryptophane concentrations typically used for hyperpolarized ¹²⁹Xe NMR biosensing experiments. Flow cytometry revealed 4-fold higher cellular internalization in cancer cells overexpressing the integrin receptor compared to normal cells. Nanomolar inhibitory concentrations (IC₅₀ = 20–30 nM) were measured for cryptophane biosensors against vitronectin binding to α_vβ₃ integrin and fibrinogen binding to α_IIbβ₃ integrin. Functionalization of the conjugate with two propionic acidgroups improved water solubility for hyperpolarized ¹²⁹Xe NMR spectroscopic studies, which revealed a single resonance at 67 ppm for the ¹²⁹Xe-cryptophane–cyclic RGDyK biosensor. Introduction of α_IIbβ₃ integrin receptor in detergent solution generated a new “ bound” ¹²⁹Xe biosensor peak that was shifted 4 ppm downfield from the “free” ¹²⁹Xe biosensor.