August 4, 2014
Caltech Team Develops Technique To Create See-Through Organs
redOrbit Staff & Wire Reports - Your Universe Online
In a discovery that could improve our general knowledge of biology, lead to more accurate clinical diagnoses and disease monitoring, and serve as a catalyst for new therapies for a variety of conditions, researchers from the California Institute of Technology (Caltech) have discovered a way to see through tissues and organs.
“Large volumes of tissue are not optically transparent – you can't see through them,” senior author Viviana Gradinaru, an assistant professor of biology at Caltech and the principal investigator in the team behind the new technique, explained in a statement.
“So, if we need to see individual cells within a large volume of tissue,” such as a tumor biopsy, “we have to slice the tissue very thin, separately image each slice with a microscope, and put all of the images back together with a computer,” she added. “It's a very time-consuming process and it is error prone, especially if you look to map long axons or sparse cell populations such as stem cells or tumor cells.”
According to Gradinaru and her colleagues, lipids are dispersed through cells and provide structural support, but they also prevent light from passing through those cells. However, the researchers have devised a way to save time by making an organism’s entire body clear, allowing them to view it in 3D using standard optical methods such as confocal microscopy (which increases resolution and contrast by eliminating out-of-focus light).
Previously, the Caltech team created a technique known as CLARITY, in which a rodent brain was infused with a solution of lipid-dissolving detergents and a water-based polymer hydrogel (which provided structural support). This made the tissues appear to be clear, but left its 3D architecture intact for study.
Building on that method, Gradinaru and her collaborators created a transparent whole-brain specimen. The new technique, perfusion-assisted agent release in situ (PARS), uses an organism’s own network of blood vessels to quickly deliver the lipid-dissolving hydrogel and chemical solution throughout the body.
Once the target area is made transparent, researchers can use standard microscopy techniques to easily look through a thick mass of tissue to focus on specific single cells that had been genetically marked with fluorescent proteins. Even without those proteins, however, the study authors said that the PARS method can be adapted to deliver stains and dyes to specific cell types when whole-body clearing is not required.
When used on an individual organ level, the technique is known as the passive clarity technique (PACT). To ensure that stripping lipids from cells did not remove other molecules, such as RNA, DNA or proteins, Gradinaru recruited Caltech colleague and assistant professor of chemistry Long Cai to verify that strands of RNA were still present and detectable with single-molecule resolution in the cells of the transparent organisms.
“Although the idea of tissue clearing has been around for a century, to our knowledge, this is the first study to perform whole-body clearing, as opposed to first extracting and then clearing organs outside the adult body,” explained Gradinaru. “Our methodology has the potential to accelerate any scientific endeavor that would benefit from whole-organism mapping, including the study of how peripheral nerves and organs can profoundly affect cognition and mental processing, and vice versa.”
“Our easy-to-use tissue clearing protocols, which employ readily available and cost-effective reagents and equipment, will make the subcellular interrogation of large tissue samples an accessible undertaking within the broader research and clinical communities,” she added.