Abstract
The development of tissue clearing technologies allows 3D imaging of whole tissues and organs, especially in studies of the central nervous system innervated throughout the body. Although the three-dimensional imaging of solvent-cleared organs (3DISCO) method provides a powerful clearing capacity and high transparency, the rapid quenching of endogenous fluorescence and peroxide removal process decreases its practicability. This study provides a modified method named tDISCO to solve these limitations. The tDISCO protocol can preserve AAV-transduced endogenous EGFP fluorescence for months and achieve high transparency in a fast and simple clearing process. In addition to the brain, tDISCO was applied to other organs and even hard bone tissue. tDISCO also enabled us to visualize the long projection neurons and axons with high resolution. This method provides a fast and simple clearing protocol for 3D visualization of the AAV- transduced long projection neurons throughout the brain and spinal cord.