Sugar solution makes tissues see-through for brain imaging at unprecedented resolution

June 25, 2013
embryo_SeeDB

(Credit: RIKEN)

RIKEN Center for Developmental biology researchers have developed a new sugar and water-based solution that turns tissues transparent in just three days, without disrupting the shape and chemical nature of the samples.

Combined with fluorescence microscopy, this technique enabled them to obtain detailed images of a mouse brain at unprecedented resolution.

Over the past few years, teams in the USA and Japan have reported a number of techniques to make biological samples transparent; they have enabled researchers to look deep down into biological structures like the brain.

“However, these techniques have limitations because they induce chemical and morphological damage to the sample and require time-consuming procedures,” explains Dr. Takeshi Imai, who led the study.

SeeDB, an aqueous fructose solution that Dr. Imai developed with colleagues Drs. Meng-Tsen Ke and Satoshi Fujimoto, overcomes these limitations.

Using SeeDB, the researchers were able to make mouse embryos and brains transparent in just three days, without damaging the fine structures of the samples, or the fluorescent dyes they had injected in them.

Callosal axon pathways (commissural fibers) imaged with SeeDB. Inset in top left corner shows the epifluorescence image of the brain, and the dashed red box indicates the area imaged. Scale bar represents 500 microns. (Credit: RIKEN)

They could then visualize the neuronal circuitry inside a mouse brain, at the whole-brain scale, under a customized fluorescence microscope without making mechanical sections through the brain.

They describe the detailed wiring patterns of commissural fibers (in the corpus callosum) connecting the right and left hemispheres of the cerebral cortex, in three dimensions, for the first time. They also report that they were able to visualize in three dimensions the wiring of mitral cells in the olfactory bulb, which is involved the detection of smells, at single-fiber resolution.

“A recently reported clearing technique, CLARITY [see "Creating a transparent brain"], is powerful and advantageous for antibody staining of thick samples, but it requires complicated procedures,” the authors say in the paper. “Because our protocol is quick, easy, inexpensive, safe and requires no special equipment, SeeDB could prove useful for a broad range of researchers working in neuroscience and developmental biology.”

UPDATE June 26, 2013 — “The SeeDB image resolution is about one micron [millionth of a meter], which is sufficient for single neuronal fiber tracing. Our method is 3D. Much higher resolution than MRI or CT scan, [equal to] electron microscopy (EM). Our method is much easier than EM, and unlike EM, our fluorescent microscopy-based approach is accessible to most researchers in biology.” — Takeshi Imai, Ph.D., Laboratory Head, Laboratory for Sensory Circuit Formation, RIKEN Center for Developmental Biology