'Light sheets' image life in 3D

Images taken from the research team's 3D microscope

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Painting a better picture of life going about its business at the microscopic scale requires a trick of the light.

A report in Nature Methods describes how "light sheets" allow researchers to take images of cellular processes in action, in unprecedented detail.

These slivers of light illuminate just the part of a living cell that is in focus, and 3D images are made from many of these thin planes stacked up.

The approach could provide a previously unachievable view of living things.

Start Quote

We have for the first time a technology that allows you to look at the three-dimensional complexity of what's going on”

End Quote Eric Betzig

That is because the very best imaging methods known so far do their work on cells that are fixed in place and whose cellular machinery has ground to a halt.

"Most of the techniques I've developed look at dead cells," said Eric Betzig, the Howard Hughes Medical Institute (HHMI) physicist who led the research.

"You can get a lot of information looking at fixed, dead cells - high-resolution information - but you'd still like to be able to see dynamics," he told BBC News.

"There's a lot you can learn from actually watching things wiggle around."

Light relief

The principal techniques in cell biologists' toolboxes are known as confocal and wide-field microscopy. But they suffer from two shortcomings, both caused by the nature of the light that is used to illuminate the sample.

One is that spatial resolution - the size down to which objects like cell components can be distinctly resolved - is not the same in all directions, leading to "elongated blobs" in images.

But another is the fact that cells do not appreciate being in the limelight for long.

(E Betzig, T Planchon, L Gao) The technique allows an array of cellular processes to be seen in action

"When you try to study live cells for any length of time, the light itself starts to harm the cells, and eventually they literally curl up and die," Dr Betzig explained.

"So there needs to be some way of getting around that."

The solution is known as plane illumination.

Instead of shining light through a sample from the bottom and looking at what passes through it, plane illumination aims to shoot light in from the side in a thin sheet, only in the plane on which a microscope is focused.

The image is formed from what bounces off the sample and up toward the microscope's lens.

This plane illumination has been used to great effect before, but the new publication takes the approach to a level of resolution both in space and in time that is unprecedented.

The secret is the use of what are known as Bessel beams (recently highlighted in a report detailing how lasers can be used as "tractor beams").

Rather than being uniform across their width, Bessel beams have a strong, narrow central point and are much weaker at the side.

The team also used what is known as a two-photon approach to ensure that the central portion of the beam - what Dr Betzig calls the "long pencil of light" - is the only part that contributes to an image.

By scanning their Bessel beams rapidly across living samples and flashing them on and off, the team could build up two-dimensional pictures as tiny strips of their sample were illuminated.

By then slightly shifting upward and downward the plane at which the microscope was focused, a number of these 2D slices could be acquired, and "stacked together" to create a 3D image.

The team can create 200 of these slices in a second, forming an image of whole, living cells - and single cell parts - caught in the act of, for instance, cell division and signalling.

While a wealth of other imaging techniques can offer higher resolution, the team's effort is superlative for the study of living cells.

They have improved the resolution through the sample - the fineness of detail they can see - by more than a factor of three over prior cell-imaging techniques, and they can acquire images far faster.

"We have for the first time a technology that allows you to look at the three-dimensional complexity of what's going on, at the sort of rates at which things happen within cells," Dr Betzig said.

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