An integrated Overhauser-enhanced MRI-Prepolarized MRI system was developed to obtain radiobiological information that could be accurately coregistered with diagnostic quality anatomic images. EPR and NMR images were acquired through the double resonance technique and field cycling of the main magnetic field from 5 mT to 0.5 T. Dedicated EPR and NMR coils were devised to minimize radiofrequency power deposition with high signal-to-noise ratio. Trityl and nitroxide radicals were used to characterize oxygen and redox sensitivities of multispin echo Overhauser-enhanced MRI. Oxygen resolution of 3 mmHg was obtained from 2 mM deoxygenated trityl phantoms. Trityl radicals were stable in reducing environments and did not alter the redox-sensitive decaying rate of the nitroxide signals. Nitroxide radicals had a compounding effect for the trityl oximetry. Tumor oxygenation and redox status were acquired with anatomical images by injecting trityl and nitroxide probes subsequently in murine tumors. The Overhauser-enhanced MRI-Prepolarized MRI system is ready for quantitative longitudinal imaging studies of tumor hypoxia and redox status as radiotherapy prognostic factors.