3-D Images Built by Mapping Negative Space with Super-Resolution Microscope

Scientist are constantly striving to study cells at a better resolution to understand their function better. Scientists at The University of Texas in Austin are now able to do this by creating three-dimensional images of structures in biological material under natural conditions. The team is hoping that this will help engineers developing artificial organs such as heart tissue or skin, while at the same time reveal how cells communicate with one another.

Using thermal noise imaging, physicist Ernst-Ludwig Florin managed to capture nanometer-scale images of networks of collagen fibrils. The fibrils are found in the skin of animals and form part of the connective tissue. A nanometer is one-hundred-thousandth of the width of a human hair or a billionth of a meter. By working on this tiny scale, scientist have for the first time managed to measure properties that affect skin’s elasticity. They envisage that this could lead to better designs for artificial skin or tissues

Biological samples are generally soft and bathed in liquid. In an effect known as Brownian motion, structures move back and forth with very small changes in temperature. This makes taking crisp 3-D images of nanoscale structures very difficult as the movement creates blurriness in the images. To overcome this problem, chemicals that stiffen various structures are often added to the sample when using super-resolution imaging techniques. This does however often cause the materials to lose their natural mechanical properties. Blurriness can also be eliminated by sticking structures to a glass structure and then focusing on that. This does however severely limit the kinds of structures and configurations that can be studied.

Florin and his team did things differently by adding nanospheres to their biological samples under natural conditions. These nanometer-sized beads reflect laser light. The scientists then shone a laser on the sample and took superfast snapshots of the nanospheres through a light microscope. This methodology is called thermal noise imaging.

Thermal noise imaging works similar to having to take a three-dimensional image of a room in total darkness. If you throw a glowing rubber ball into the room and take a series of high-speed images of the ball as it bounces around, you would see that as the ball bounces around the room, it can only move in the unoccupied spaces. Taking millions of images so fast that they don’t blur and combining them, would show an overall picture of where there are objects. Solid objects such as tables and chairs would show up as blanks on the picture.

The equivalent of the rubber ball in thermal noise imaging is a nanosphere that moves around in a sample by natural Brownian motion. Florin explains that while the chaotic movement is an annoyance for most microscopy techniques because it makes everything blurry, they use the same motion to their advantage. Nature literally does the work for them without any need for complicated mechanisms to move a probe around.

The thermal noise imaging technique was already published and patented in 2001. It has however not been developed into a fully functioning process due to technical challenges.

Researchers have now measured the mechanical properties of collagen fibrils in a network for the first time. Collagen is a biopolymer that contributes to the skin’s elasticity by forming frameworks for cells in the skin. An important question that must be answered by scientists before the rational design of artificial skin can proceed, is how a collagen network’s architecture translates into its elasticity. Florin notes that once this knowledge is available, better collagen networks could be designed that act as a framework to encourage cells to grow correctly.

The research is fully described in the journal Nature Communications.