Mirror image: Researchers create higher-quality pictures of biospecimens

November 13, 2017, National Institutes of Health
The images obtained by the combination of the new coverslip and computer algorithms show clearer views of small structures. Credit: Yicong Wu, National Institute of Biomedical Imaging and Bioengineering

Researchers from the National Institutes of Health and the University of Chicago improved the speed, resolution, and light efficiency of an optical microscope by switching from a conventional glass coverslip to a reflective, mirrored coverslip and applying new computer algorithms to process the resulting data.

Hari Shroff, Ph.D., chief of the National Institute of Biomedical Imaging and Bioengineering's lab section on High Resolution Optical Imaging (HROI), and his team have spent the last few years developing optical microscopes that produce high resolution images at very high speed. After his lab develops these new microscopes, they release the plans and software for free, so any researcher can replicate the advances made at NIH.

This latest microscope builds on previous improvements that Shroff's lab had made with selective plane illumination microscopy (SPIM). The developments are described in a paper published Nov. 13, 2017, in the advance online edition of Nature Communications. SPIM systems differ from traditional microscopes because they use sheets to excite the sample, only exposing the imaged sample plane to light. Because only the portion of the sample being imaged (rather than the entire sample) is exposed to light, there is less overall damage to the sample. Thus, SPIM systems are gentler than traditional microscopes.

In 2013, Shroff and his colleague in the HROI lab, Yicong Wu, developed the diSPIM—a SPIM system equipped with two lenses so it obtains two views of the sample instead of just one. Just as using two eyes provides much better depth and three-dimensional perception than using only one eye, the dual view microscope enables 3-D imaging with much greater clarity and resolution than traditional single-view imaging. In 2016, they added a third lens, showing that this additional view can further improve light efficiency and resolution in 3-D imaging.

"Once we incorporated three lenses, we found it became increasingly difficult to add more," said Shroff. "Not because we reached the limit of our computational abilities, but because we ran out of physical space."

Mirror image: Researchers create higher-quality pictures of biospecimens
In this diagram, you can see how the mirrored coverslip allows for four simultaneous views. Credit: Yicong Wu, National Institute of Biomedical Imaging and Bioengineering

The lenses used to image the samples are bulky and need to be close to the samples to clearly image the detailed subcellular structure within a single cell or the neuronal development within a worm embryo. The space around the sample becomes more and more limited with each additional .

Wu and Shroff's solution was conceptually simple and relatively low-cost. Instead of trying to find ways to stuff in more lenses, they use mirrored coverslips.

"It's a lot like looking into a mirror," Shroff explained. "If you look at a scene in a mirror, you can view perspectives that are otherwise hidden. We used this same principle with the microscope. We can see the sample conventionally using the usual views enabled by the lenses themselves, while at the same time recording the reflected images of the sample provided by the mirror."

One complication is that both the conventional and reflected views contain an unwanted background generated by the light source. In order to deal with this problem, Wu and Shroff collaborated closely with Patrick La Riviere's group at the University of Chicago. La Riviere is an expert in computational imaging, and helped the team create computer processing software that can identify and remove the unwanted background and clarify the image.

Using the mirrored coverslips in conjunction with the computer software, the team was able to improve the speed two-fold and almost double the resolution in comparison with conventional diSPIM systems without changing the hardware of the microscope. An additional benefit of the technique is that with mirrored coverslips, the is able to collect more light from the sample without increasing the overall light exposure to the . As a result, it increases the efficiency by two to three times compared with diSPIM. The researchers hope that in the future this technique may be adapted to other forms of microscopy.

Explore further: New approach doubles 3-D resolution of fluorescence microscopy

More information: Yicong Wu et al, Reflective imaging improves spatiotemporal resolution and collection efficiency in light sheet microscopy, Nature Communications (2017). DOI: 10.1038/s41467-017-01250-8

Related Stories

A microscope within a microscope

August 14, 2017

No single microscope can image all aspects of a sample at the same time and so the use of two or more imaging methods to study a sample - correlative imaging - is common-place.

Microfabrication leads to a new microscopy method

June 18, 2015

With the aim of providing a simple, yet versatile microscopy technique that can identify single proteins anywhere within a cell, and allow cellular organization to be assessed in 3D, researchers at the Mechanobiology Institute ...

New Microscope Gives Scientists 3D Views of Living Organisms

August 12, 2004

Physicists at the European Molecular Biology Laboratory (EMBL) have developed a state-of-the-art microscope that gives scientists a much deeper look into living organisms than ever before. The new technology will undoubtedly ...

Pushing microscopy beyond standard limits

July 29, 2013

Engineers at the California Institute of Technology (Caltech) have devised a method to convert a relatively inexpensive conventional microscope into a billion-pixel imaging system that significantly outperforms the best available ...

Recommended for you

ATLAS experiment observes light scattering off light

March 20, 2019

Light-by-light scattering is a very rare phenomenon in which two photons interact, producing another pair of photons. This process was among the earliest predictions of quantum electrodynamics (QED), the quantum theory of ...

How heavy elements come about in the universe

March 19, 2019

Heavy elements are produced during stellar explosion or on the surfaces of neutron stars through the capture of hydrogen nuclei (protons). This occurs at extremely high temperatures, but at relatively low energies. An international ...

Trembling aspen leaves could save future Mars rovers

March 18, 2019

Researchers at the University of Warwick have been inspired by the unique movement of trembling aspen leaves, to devise an energy harvesting mechanism that could power weather sensors in hostile environments and could even ...

Quantum sensing method measures minuscule magnetic fields

March 15, 2019

A new way of measuring atomic-scale magnetic fields with great precision, not only up and down but sideways as well, has been developed by researchers at MIT. The new tool could be useful in applications as diverse as mapping ...


Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.