Molecular imaging provides the ability to study cellular and molecular processes that have the potential to impact many facets of biomedical research. Fluorescence imaging, one technique of molecular imaging, is a technique which has been widely used. Nitric oxide and peroxynitrite are important signaling molecules in organism. They are involved in diverse physiological and pathological pathway. Monitoring and detecting the level and distribution of nitric oxide and peroxynitrite by fluorescent imaging will be helpful in the understanding of roles played by these molecules. In this review, we introduce the fluorescent probes of nitric oxide and peroxynitrite with their design strategies and development.