The adage “Seeing is believing” was on the minds of Dr. Chih-Wei Logan Hsu and Dr. Joshua D. Wythe of Baylor College of Medicine as they and their colleagues developed an innovative technology called EZ Clear. This new method of tissue clearing has simplified and accelerated the process of making tissue optically transparent, allowing 3D imaging of whole intact tissues or even whole organs.
Their new method, published in the journal eLife, was developed at Baylor’s Optical Imaging and Vital Microscopy Core (OiVM). Tissue clearing and whole-organ imaging have revolutionized biology, allowing the exploration of organs in three-dimensional space without compromising tissue architecture.
“Previous methods were complicated, laborious and often required expensive equipment, as well as the use of hazardous organic solvents, which prevented widespread adoption of these methods,” said Hsu, co-director of OiVM and assistant professor of integrative physiology and education, innovation and technology at Baylor. “These challenges motivated us to develop a simpler clearing process that users could more easily complete, saving valuable time and resources to focus on the real issues they want to investigate in their systems.”
“The beauty of this method is that you can analyze the sample from an aggregate or macro perspective without physically disrupting the natural organization of the tissue or organ,” said Wythe, associate professor of integrative physiology and neurosurgery at Baylor. He is also a member of the Dan L Duncan Comprehensive Cancer Center and the Cardiovascular Research Institute.
“For example, researchers can now visualize neural connections between the eye and the brain. If done in sections, the process would disrupt natural tissue organization and is incredibly difficult to reconstruct in 3D, limiting our understanding of connections between neurons and other surrounding cells over larger volumes or areas.3D imaging circumvents these limitations, and the advent of EZ Clear makes 3D imaging accessible to most modern molecular biology labs “said Wythe.
EZ Clear also preserves endogenous and synthetic labeling methods, such as fluorescence, without changing sample size. The study shows successful clearing and labeling of neurons and blood vessels in the brain, as well as vessels in the eyes, heart, kidneys, testicles and ovaries, and successful clearing of whole organs in the lungs, mouse liver and pancreas.
“EZ Clear eliminates previous technical barriers, allowing researchers to inspect their organs of interest from a macro, whole-organ level, down to cellular resolution,” Wythe said. “It has eliminated previous practical, safety and economic challenges, while providing reproducible, high-quality visualization of the whole organ, which is an important prospect as it can provide new insights into the subject being studied.”
EZ Clear is faster, cheaper and easier than previous clearing methods. “Researchers can now teach OiVM this tissue removal process that is easier to perform and reproducible. They can then implement it in their own labs and get results in 48 hours by following three simple steps, when other methods take weeks or even months. clean the fabric,” Hsu said. “Then they can bring the cleared organ to the OiVM for 3D imaging and analysis.”
Other contributors to this work include Juan Cerda, Jason M. Kirk, Williamson D. Turner, Tara L. Rasmussen, Carlos P. Flores Suarez, and Mary E. Dickinson. The authors are all affiliated with Baylor College of Medicine.
This project was supported by the Optical Imaging and Vital Microscopy Core and the Bioengineering Core at Baylor College of Medicine. Additional support was provided by grants from the National Institutes of Health (5T32GM088129-10, R01HL146745, R01HD099026, U42OD026645, R01HL159159, and 1S10OD016167), American Heart Association (22PRE916015), Cancer Prevention Research Institute of Texas (RP200402) , the Department of Defense (W81XWH18-1-0350) and the Canadian Institutes of Health Research (PJT-155922).