Preclinical translational imaging, and specifically molecular imaging methods such as positron emission tomography (PET) or single photon emission computed tomography (SPECT) as well as magnetic resonance imaging (MRI) are extremely useful tools in the drug discovery and development process. Continuous progress in modern imaging technologies holds a great potential to extend efficiencies to drug discovery and translation and to reduce clinical attrition. Newer technologies such as photoacoustic tomography and confocal laser endomicroscopy lend the ability to identify critical and highly specific biomarker information for oncology, neurology, and immunology strategies. These approaches can significantly enhance preclinical and translational research. The discussion will provide an overview of the strengths of different imaging technologies and will focus on scientific translational applications in oncology, neurology, immunology, and safety.