proteins in a cell

Proteins within a cell, seen with techniques developed by Eric Betzig. Image courtesy HHMI

For figuring out a way to see things that most scientists thought could never be seen, three microscope specialists have won the 2014 Nobel Prize in chemistry. Stefan W. Hell, and Eric Betzig with William E. Moerner, developed two different techniques for increasing the power of light microscopy, allowing scientists to see molecules in action within a living cell, watch DNA being put together, and follow the actions of proteins involved in Huntington’s disease and Alzheimer’s disease.

This morning in Stockholm, the Nobel committee announced that the prize, “for the development of super-resolved fluorescence microscopy,” was being given to Hell, a director at the Max Planck Institute for Biophysical Chemistry in Göttingen, Germany; Betzig, a researcher at the Janelia Farm Research Campus of the Howard Hughes Medical Institute in Ashburn, Virginia; and Moerner, a professor at Stanford University in California.

The scientists figured out ways to break a seemingly untouchable barrier laid down in 1873, when microscopist Ernst Abbe wrote an equation setting the bottom limit of the light microscope. His work indicated, very persuasively, that the visible world stopped at about half the wavelength of visible light, or about 0.2 micrometers. This “diffraction limit” means that scientists could see the outlines of a cell, but not its inner workings.

Electron microscopes could go smaller, said Sven Lidin, chair of the Nobel chemistry committee and a chemist at Lund University in Sweden. “But using them meant you had to kill the cell,” which made it impossible to see anything in action, he said. The new Nobelists' work means that "reactions can be studied as they happen, not as the end result but actually as they take place. It opens entirely new possibilities for chemistry and for biochemistry.”

Hell, in 1994, published an article describing a method called stimulated emission depletion that, in essence, turned an optical microscope into a flashlight, able to focus on a very small part of a cell. The method used two pulses of light. The first pulse stimulates fluorescent molecules in a cell. The second, “quenching,” pulse cancels out the light from all the molecules that surround a single, glowing point. This allows the glowing area to be seen clearly. In 2000, Hell was able to image a single bacterium at a resolution never before achieved with an optical microscope.

Betzig and Moerner, independently, developed another technique called single molecule microscopy. The idea is to selectively turn on some fluorescent molecules in a sample, but leave others dark. Then move the focus slightly to adjacent molecules, and repeat. In 1997 Moerner showed he could home in on just a single molecule, green fluorescent protein from a jellyfish, making it light up and go dark like a tiny lamp with a switch. And in 2006, Betzig showed that after one molecule went dark, he could light up one nearby, and by superimposing the images, create a complete picture of part of a cell.

Reached by telephone this morning, Hell said the diffraction barrier was a daunting one. “But I was confident that turning molecules on and off could show the way. I didn’t give up,” he said. Lidin, from the Nobel committee, added that to breach such a barrier, “you have to have a lot of confidence in your ideas. And you need stamina.”

Tom Barton, president of the American Chemical Society and a professor at Iowa State University, said the winners’ work “allowed us the see the previously unseen—lifting the veil on bacteria, viruses, proteins, and small molecules.”

Betzig has built on these techniques to make high-resolution movies of cells, and you can read about them (and see the movies) in Scientific American, which described this work in 2013.

Moerner's method for looking inside cells was described in a 2009 Scientific American article. His technique for analyzing single molecules within a crystal was reported on by the magazine in 1991.