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This paper presents a novel approach for 3D imaging of the mouse brain in vivo at high magnetic field. High resolution 3D in vivo mouse brain images provide important information for understanding the relationship between genotype and phenotype in transgenic mice models, which are widely used in translational research of human diseases. Compared to ex vivo, in vivo imaging is more accurate and essential for longitudinal studies. High resolution MRI of a mouse brain in vivo is very challenging because the quality of the image is impacted by animal motion, physiological noise, MRI susceptibility and scan time limitation. Through our efforts to optimize the hardware, anesthesia technique, animal monitoring and MRI sequence design, we have achieved high quality 3D in vivo images at 100mum resolution, which allows for future accurate morphological studies.