In vivo optical imaging is a technology that allows visualization of the inside of an animal's body while it is still alive by illuminating only specific cells (e.g., cancer). We have been working to develop new materials based on luciferin, a bioluminescent substance found in fireflies. Based on this research, we have successfully synthesized a luminescence substrate with a luminescence peak in the window region of the living body (650 nm to 900 nm). This makes it possible to observe living organisms by making them glow while they are alive and enables imaging down to a depth of 5 to 6 cm from the epidermis. Furthermore, Aka-BLI, a combination of Tokeoni optimized mutant “AkaLuc” and Tokeoni, enables quantitative observation of a small number of cells that have been difficult to detect. In addition, Aka-BLI can be used to quantitatively observe neurons in the brain. Aka-BLI is 1000 times stronger than green-BLI, which combines firefly luciferin and wild-type luciferase, and also successfully used for imaging of marmoset striatum.