Label-free photoacoustic microscopy of cytochrome c in cells

Cytochrome c is a heme protein normally bound to mitochondria and is important for mitochondrial electron transport and apoptosis initiation. Since cytochrome c is nonfluorescent, it is always labeled with fluorescent molecules for imaging, which, however, may affect normal cellular functions. Here, label-free photoacoustic microscopy (PAM) of mitochondrial cytochrome c was realized for the first time by utilizing the optical absorption around the Soret peak. PAM was demonstrated to be sensitive enough to image mitochondrial cytochrome c at 422 nm wavelength. Mitochondrial cytochrome c in the cytoplasm of fixed fibroblasts was clearly imaged by PAM as confirmed by fluorescent labeling. By showing mitochondrial cytochrome c in various cells, we demonstrated the feasibility of PAM for label-free histology of mouse ear sections. Therefore, PAM can sensitively image cytochrome c in unstained cells at 422 nm wavelength and has great potential for functional imaging of cytochrome c in live cells or in vivo.