NIfTI-Studio is a Matlab toolbox enabling researchers to visualize 3D NIfTI and Analyze images. Users can flip through slices, change orientations, 3D render surfaces, plot mosaics of slices, or add statistical overlays. NIfTI-Studio also allows for the manual creation of regions of interest (ROIs) to support additional analyses. The toolbox features an intuitive user interface that is easy to learn, but users also have the option to perform nearly all tasks via command line calls. The toolbox was designed for use with MRI brain imaging data, but it should be compatible with any NIfTI or Analyze format images.