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The aim of this study was to test and demonstrate the feasibility of diffusion tensor imaging (DTI) with a hybrid positron emission tomography (PET)/magnetic resonance imaging system for simultaneous PET and magnetic resonance (MR) data acquisition.All measurements were performed with a prototype hybrid PET/MR scanner dedicated for brain and head imaging. The PET scanner, which is inserted into a conventional 3.0-Tesla high field MR imager equipped with a transmit/receive birdcage head coil, consists of 192 block detectors with a matrix of 12 × 12 lutetium oxyorthosilicate scintillation crystals combined with MR-compatible 3 × 3 avalanche photodiode arrays. In 7 volunteers and 4 patients with brain tumors, DTI was performed during simultaneous PET data readout applying a diffusion weighted echo planar sequence (12 noncollinear directions, echo time (TE)/repetition time (TR) 98 ms/5300 ms, b-value 800 s/mm2). Image quality and accuracy of DTI were assessed in comparison with DTI images acquired after removal of the PET insert.The diffusion images showed good image quality in all volunteers regardless of simultaneous PET data readout or after removal of the PET scanner; however, significantly (P < 0.01) stronger rim artifacts were found in fractional anisotropy images computed from DTI images recorded during simultaneous PET acquisition, demonstrating higher eddy-current effects. In region of interest analysis, no notable differences were found in the computation of the direction of the principal eigenvector (P > 0.05) and fractional anisotropy values (P > 0.05). In the assessment of pathologies, in all 4 patients PET and DTI provided important clinical information in addition to conventional magnetic resonance imaging.Diffusion tensor imaging may be combined with simultaneous PET data acquisition, offering additional important morphologic and functional information for treatment planning in patients with brain tumors.