Merging old and new microscopy yields best 3D view yet inside cells

Cells are filled with small vesicles: membrane-bound sacks that help cells store proteins, break down cellular garbage and carry cargo. These many varieties of vesicles are indistinguishable from each other under an electron microscope, but with cryo-SR/EM, their distinct features become clear. This clip zooms in on endosomes, which shuttle cargo to different regions within the cell. (Video courtesy of D. Hoffman et al, HHMI)

By combining electron microscopy (EM) with the latest super-resolution microscopy (SR), Howard Hughes Medical Institute (HHMI) and UC Berkeley scientists have obtained exquisitely detailed views of the complex innards of cells, all in 3D.

In a report in this week’s issue of the journal Science, the researchers describe their technique, called cryo-SR/EM, and display some of the colorful images they captured. These include detailed pictures of the tentacled vesicles that shuttle and sort cargo in the cell, views of a delicate web that connects neighboring neurons in the brain, and images of the rearrangement of DNA in the nucleus as a stem cell differentiates into a neuron.

“The cell is an incredibly complex self-replicating machine with billions of moving parts: molecules. Each imaging technique sees this machine in different, yet incomplete, ways,” said co-author Eric Betzig, a UC Berkeley professor of molecular and cell biology and of physics and a Howard Hughes Medical Institute investigator. “Bringing two very different imaging modalities together, as we have with cryo-SR and EM, brings insights into cellular structure at the nanoscale, which cannot be obtained by either alone.”

Light microscopy, for example, makes it simple to identify specific cellular structures by tagging them with easy-to-see fluorescent molecules. With the development by Betzig and others of super-resolution (SR) fluorescence microscopy, these structures can be viewed with even greater clarity. But fluorescence can reveal only a few of the more than 10,000 protein molecules in a cell at a given time, making it difficult to understand how these few relate to everything else.

Electron microscopy, on the other hand, reveals all cellular structures in high-resolution pictures, but delineating one feature from all others by EM alone can be difficult because the space inside of cells is so crowded.

Combining the two techniques gives scientists a clear picture of how specific cellular features relate to their surroundings, said Harald Hess, a senior group leader at HHMI’s Janelia Research Campus. “This is a very powerful method.”

Janelia research scientist David Hoffman and senior scientist Gleb Shtengel spearheaded the project under the leadership of Hess and Betzig, who is also a Janelia senior fellow.

New microscopy technique shows cells’ 3D ultrastructure in new detail