Visualising cell structures in three dimensions

Viral pathogens like the SARS-CoV-2 coronavirus change the interior structure of the cells they infect, requiring the use of extremely powerful imaging techniques to visualise these changes. A German–American research team, working under the direction of Dr Venera Weinhardt at Heidelberg University’s Centre for Organismal Studies (COS), recently optimised a special X-ray process — known as soft X-ray tomography — to deliver high-resolution three-dimensional images of entire cells and their molecular structure in just a few minutes. The team’s results were published in the journal Cell Reports Methods.

“Scanning electron microscopes are preferred in cell imaging because they provide extremely sharp nanoscale images,” said Dr Weinhardt, a postdoc at the COS and at Lawrence Berkeley National Laboratory in the US. “But this technology takes a good week to scan an individual cell. It also generates an enormous amount of data that is daunting to analyse and interpret. Using soft X-ray tomography, we get usable results within 5–10 minutes.”

High throughput is extremely important for studying numerous cells, according to molecular virologist Prof Dr Ralf Bartenschlager, whose department at Heidelberg University Hospital is collaborating with Dr Weinhardt on imaging cellular changes associated with viral infections. In tissue, Prof Dr Bartenschlager said, often only some of the cells are infected. Only those cells provide information on the changes that result directly from the infection. Looking for these cells with a scanning electron microscope, however, is not possible.

Soft X-ray tomography (SXT) has already been used to successfully detect single virus particles — called virions — of different types of viruses and their associated changes in cells. Now the researchers have used the technology to study cell cultures infected with SARS-CoV-2 from lung and kidney tissue. Soft X-rays allowed them to image complete cells and their structure in three dimensions in 5–10 minutes. The researchers were further able to detect clusters of SARS-CoV-2 particles on cell surfaces as well as identify virus-associated changes in the cell’s interior. Structures were revealed that possibly enable the replication and spread of the virus.

According to Dr Weinhardt, the team’s success largely hinged on the technology allowing them to study fixed cells, ie, cells that had been chemically treated to deactivate the virus. Typically, in soft X-ray tomography, like in electron tomography, flat lattice structures are used as holders. When they are tilted, the thickness of the samples can change, making some cell structures appear blurry. ‘Blind’ spots also occur because the flat shape of the holder prevents the cells from being scanned at all angles. Another dilemma is that the samples can adhere to the lattice or spread out, requiring multiple tomograms to visualise the entire cell.

“To get around this problem, we switched over to cylindrical thin-wall glass capillaries to hold the samples,” Dr Weinhardt said. “During microscopy, the samples can be rotated a full 360 degrees and scanned from all angles.”

The team is now working on further refining sample preparation techniques, automating the analysis of the 3D image data, and developing a laboratory version of a soft X-ray microscope.

Image caption: Human lung epithelium cell 24 hours after SARS-CoV-2 viral infection. Hijacked cellular organelles are labelled with an asterisk. Image ©Venera Weinhardt (COS).

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