RTF: a rapid and versatile tissue optical clearing method

Tissue optical clearing enables imaging deeper in large volumes with high-resolution. Clear T2 is a relatively rapid clearing method with no use of solvents or detergents, hence poses great advantage on preservation of diverse fluorescent labels. However, this method suffers from insufficient tissue transparency, especially for adult mouse brain blocks. In this work, we develop a rapid and versatile clearing method based on Clear T2 , termed RTF (Rapid clearing method based on Triethanolamine and Formamide), aiming for better clearing capability. The results show that RTF can not only efficiently clear embryos, neonatal brains and adult brain blocks, but also preserve fluorescent signal of both endogenous fluorescent proteins and lipophilic dyes, and be compatible with virus labeling and immunostaining. With the good transparency and versatile compatibility, RTF allows visualization and tracing of fluorescent labeling cells and neuronal axons combined with different imaging techniques, showing potentials in facilitating observation of morphological architecture and visualization of neuronal networks.