Efficient 3D light-sheet imaging of very large-scale optically cleared human brain and prostate tissue samples

The ability to image human tissue samples in 3D, with both cellular resolution and a large field of view (FOVs), can improve fundamental and clinical investigations. Here, we demonstrate the feasibility of light-sheet imaging of ∼5 cm3 sized formalin fixed human brain and up to ∼7 cm3 sized formalin fixed paraffin embedded (FFPE) prostate cancer samples, processed with the FFPE-MASH protocol. We present a light-sheet microscopy prototype, the cleared-tissue dual view Selective Plane Illumination Microscope (ct-dSPIM), capable of fast, 3D high-resolution acquisitions, of cubic centimetre sized cleared tissue. We used Mosaic scans for fast 3D overview scans of entire tissue samples or higher resolution overviews of large ROIs with various speeds: a) Mosaic 16 (16.4 µm isotropic resolution, ∼ 1.7 hr/cm3), b) Mosaic 4 (4.1 µm isotropic resolution, ∼ 5 hr/cm3) and c) Mosaic 0.5 (0.5 µm near isotropic resolution, ∼15.8 hr/cm3). We could visualise ROIs around the border of human brain area V1/V2, and could demonstrate suitable imaging quality for Gleason score grading in prostate cancer samples. We show that ct-dSPIM imaging is an excellent technique to quantitatively assess entire MASH prepared large-scale human tissue samples in 3D, with considerable future clinical potential in prostate cancer.