Efficient optical clearance is fundamental for whole brain imaging. In particular, clearance of the brain without membrane damage is required for the imaging of lipophilic tracer-labeled neural tracts. Relying on an ascending gradient of fructose solutions, SeeDB can achieve sufficient transparency of the mouse brain while ensuring that the plasma membrane remains intact. However, it is challenging to extend this method to larger mammalian brains due to the extremely high viscosity of the saturated fructose solution. Here we report a SeeDB-derived optical clearing method, termed FRUIT, which utilizes a cocktail of fructose and urea. As demonstrated in the adult mouse brain, combination of these two highly water-soluble clearing agents exerts a synergistic effect on clearance. More importantly, the final FRUIT solution has low viscosity so as to produce transparency of the whole adult rabbit brain via arterial perfusion, which is impossible to achieve with a saturated fructose solution. In addition to good compatibility with enhanced yellow fluorescent protein, the cocktail also preserves the fluorescence of the lipophilic tracer DiI. This work provides a volume-independent optical clearing method which retains the advantages of SeeDB, particularly compatibility with lipophilic tracers.
Ke Yang,Xiang Yang,Xiang Zhao,Marc Lamy de la Chapelle,Weiling Fu
Simple, rapid, and efficient cell viability assays play a fundamental role in much of biomedical research, including cell toxicology investigations and antitumor drug screening. Here, we demonstrate for the first time a rapid and label-free cell viability assay using THz spectroscopy in combination with a new optical clearing agent (OCA) and microfluidic technology. This strategy uses a considerably less absorptive OCA to replace the highly absorptive water molecules around the living cells and thus to decrease the background signal interference. Three low-viscosity oils were screened as potential OCA candidates, among which fluorinated oil was selected because of its lower absorption and lowest cytotoxicity. After the liquid medium was replaced with fluorinated oil in a microfluidic chip, an obvious THz spectral difference was observed between the fluorinated oils with and without living cells. This change in THz response was preliminarily attributed to the distinguishable signals between the cells and the fluorinated oil. In addition, we applied this method to cell viability assays of human breast cancer cells (MDA-MB-231) after treatment with different antitumor drugs. The results indicated that THz spectroscopy with the aid of the proposed water-replacement strategy presented excellent quantification of cell viability with the advantages of a rapid, label-free, nondestructive microassay, which offers significant potential to developing a convenient and practical cell analysis platform.
危险品化学品经营许可证（带存储）