New Electron Microscopy Tech Breaks into the Elusive Realm of Small Molecules

Adapted from an article by Robert Sanders at UC Berkeley.

Nearly 100 years ago, a discovery revolutionized light microscopy. The introduction of the phase-contrast microscope, which garnered a Nobel Prize in 1953, brought into clear view structures inside cells that had previously been too faint or washed out for biologists to study.

A team of physicists from UC Berkeley and Lawrence Berkeley National Laboratory (Berkeley Lab), have now adapted the phase-contrast technique to cryo-electron microscopy (cryo-EM), which has about 10,000 times the magnification of light microscopy. As reported in Science, their laser-based phase plate produces sharp images of molecules that today's cutting-edge cryo-EM systems struggle to capture.

The new technology was brought to fruition by more than 15 years of theoretical and experimental work by leading microscopy scientists, collaboration with expert machinists, and support from Biohub. The phase plate is paired with a new, custom Thermo Fisher Scientific microscope that was developed to maximize the benefit of the plate's ultra-bright laser. Images taken by the system are notably clearer and sharper and contain greater detail that structure-solving software can process to generate more accurate atomic models of the molecules captured.

"Theia is the Formula 1 of microscopes. It has better resolution than the standard cryo-EM, even without the laser. With the addition of the laser phase plate, we hope that it really becomes one of the world's best instruments overall," said Holger Müller, a UC Berkeley professor of physics and faculty scientist in Berkeley Lab's Biosciences Area who led the development effort. "Before, studying structures with cryo-EM was like trying to look at paintings in a dark gallery. With Theia, it's like the lights have been turned on for the first time."

In their paper, Müller and his colleagues demonstrate the system's power by imaging aldolase, a protein in muscle that is relatively easy to capture with today's cryo-EM machines, and hemoglobin - a protein that carries oxygen in blood. Hemoglobin is a smaller protein that sits at the lower size limit for current machines and is often used as a benchmark for cryo-EM performance. The laser-phase plate improved the resolution of the protein structure in both cases, but more so for the hemoglobin. Their study also tested the microscope on samples of varying quality to demonstrate how the phase plate can alleviate the difficult and time-intensive process of sample preparation. Currently, single-particle analyses like these require thousands to millions of isolated target molecules that have been carefully frozen to -160 degrees C or below. And even the smallest amount of ice on samples can interfere with imaging.