In vivo autofluorescence in the biological windows: the role of pigmentation

Small animal deep-tissue fluorescence imaging in the second Biological Window (II-BW, 1000-1350 nm) is limited by the presence of undesirable infrared-excited, infrared-emitted (900-1700 nm) autofluorescence whose origin, spectral properties and dependence on strains is still unknown. In this work, the infrared autofluorescence and laser-induced whole body heating of five different mouse strains with distinct coat colors (black, grey, agouti, white and nude) has been systematically investigated. While neither the spectral properties nor the magnitude of organ autofluorescence vary significantly between mouse strains, the coat color has been found to strongly determine both the autofluorescence intensity as well as the laser-induced whole body heating. Results included in this work reveal mouse strain as a critical parameter that has to be seriously considered in the design and performance of small animal imaging experiments based on infrared-emitting fluorescent markers.