The aim of this study was the evaluation of 68Ga-DOTA-E-[c(RGDfK)]2 as a novel PET tracer to image αvβ3 and αvβ5 integrins. For this purpose, DOTA-E-[c(RGDfK)]2 was labeled with 68Ga, which was obtained from a 68Ge/68Ga generator, purified by solid-phase extraction and the radiochemical purity analyzed by radio-RP-HPLC. 68Ga-DOTA-E-[c(RGDfK)]2 was obtained reproducibly in radiochemical yields of 60 ± 6% and with an excellent radiochemical purity of >99%. In nude rats bearing bone metastases after injection of MDA-MB-231 human breast cancer cells, biodistribution studies were performed to evaluate the accumulation of the radiotracer in selected organs, blood and bone metastases 0.5, 1, 2 and 3 h post injection. A rapid uptake into the bone metastases and rapid blood clearance was observed, resulting in tumor–blood ratios of up to 26.6 (3 h post injection) and tumor–muscle ratios of up to 7.9 (3 h post injection). A blocking experiment with coinjected αvβ3/αvβ5 antagonist showed the tumor uptake to be receptor-specific. In an initial in vivo micro PET evaluation of the tracer using the same animal model, the bone metastasis was clearly visualized. These results suggest that 68Ga-DOTA-E-[c(RGDfK)]2 is a promising PET tracer suitable for the imaging of αvβ3 and αvβ5 integrins in bone metastases. This novel PET tracer should be further evaluated concerning its usefulness for early detection of bone metastases and monitoring treatment response of these lesions. Copyright © 2011 John Wiley & Sons, Ltd.