An optical tissue clearing protocol for light microscopy sample preparation

Aim: To undertake an optical tissue clearing protocol to render tissue transparent enabling deep fluorescent imaging into large tissue volumes using available light microscopy technologies at Westmead Imaging, i.e. confocal and light sheet imaging.

Methods: Different tissues were optically cleared using a protocol combining multiple clearing techniques including lipid-removal, bleaching, and RapiClear 1.52. The cleared tissueswere then stained for imaging with confocal microscopy.

Results: Transparent tissues were obtained at the whole-mount scale that enabled imaging through 300um using confocal microscopy.

Conclusions: An optical tissue clearing protocol was performed using combined clearing techniques for whole-mount tissue. The protocol may be adapted to other large volume tissues to ultimately improve imaging depth using optical microscopy that is limited by the scattering of light by opaque tissues.