Figure 2.

Logic flow of the EZ Clear protocol

Delipidation times may vary depending on the age, genotype and/or disease state. Since lightsheet imaging is time-consuming, the first quality check should be verifying optical transparency using the grid paper and transmitted phase microscopy. Once optical transparency is achieved, the ultimate quality control is determined by lightsheet imaging. We recommend additional delipidation after re-equilibrating in 1X PBS for 8–12 h to improve the lightsheet imaging quality.