Three-dimensional imaging of solvent-cleared organs using 3DISCO.

Nature protocols (2012-10-13)
Ali Ertürk, Klaus Becker, Nina Jährling, Christoph P Mauch, Caroline D Hojer, Jackson G Egen, Farida Hellal, Frank Bradke, Morgan Sheng, Hans-Ulrich Dodt
ABSTRACT

The examination of tissue histology by light microscopy is a fundamental tool for investigating the structure and function of organs under normal and disease states. Many current techniques for tissue sectioning, imaging and analysis are time-consuming, and they present major limitations for 3D tissue reconstruction. The introduction of methods to achieve the optical clearing and subsequent light-sheet laser scanning of entire transparent organs without sectioning represents a major advance in the field. We recently developed a highly reproducible and versatile clearing procedure called 3D imaging of solvent-cleared organs, or 3DISCO, which is applicable to diverse tissues including brain, spinal cord, immune organs and tumors. Here we describe a detailed protocol for performing 3DISCO and present its application to various microscopy techniques, including example results from various mouse tissues. The tissue clearing takes as little as 3 h, and imaging can be completed in ∼45 min. 3DISCO is a powerful technique that offers 3D histological views of tissues in a fraction of the time and labor required to complete standard histology studies.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Tetrahydrofuran, anhydrous, ≥99.9%, inhibitor-free
Sigma-Aldrich
Tetrahydrofuran, inhibitor-free, suitable for HPLC, ≥99.9%
Sigma-Aldrich
Tetrahydrofuran, anhydrous, contains 250 ppm BHT as inhibitor, ≥99.9%
Sigma-Aldrich
Tetrahydrofuran, contains 250 ppm BHT as inhibitor, ACS reagent, ≥99.0%
Sigma-Aldrich
Tetrahydrofuran, contains 250 ppm BHT as inhibitor, puriss. p.a., ACS reagent, reag. Ph. Eur., ≥99.9%
Sigma-Aldrich
Benzyl ether, 98%
Sigma-Aldrich
Tetrahydrofuran, suitable for HPLC, ≥99.9%, inhibitor-free
Sigma-Aldrich
Tetrahydrofuran, ReagentPlus®, ≥99.0%, contains 250 ppm BHT as inhibitor
Sigma-Aldrich
Dibenzyl ether, purum, ≥98.0% (GC)
Supelco
Tetrahydrofuran, analytical standard
Supelco
Tetrahydrofuran, Selectophore, ≥99.5%
Sigma-Aldrich
Tetrahydrofuran, ACS reagent, ≥99.0%, contains 250 ppm BHT as inhibitor
Sigma-Aldrich
Tetrahydrofuran, contains 250 ppm BHT as inhibitor, ACS reagent, ≥99.0%
Sigma-Aldrich
Benzyl ether, ≥98%, FCC, FG