Microscope captures ‘lost’ fluorescence to improve resolution

Macrophage actin labeled with green fluorescent protein, imaged with the dual-view microscope (left) and with the new triple-view microscope (right).
Courtesy of
Yicong Wu and Valentin Jaumouille
Steve Koppes
Associate News DirectorUniversity Communications

Taking a cue from medical imaging, scientists have invented a multi-view microscope that captures higher-resolution, 3-D images of live cells and tissues without upping the dose of potentially harmful radiation the specimens receive.

The researchers, who work collaboratively at the Marine Biological Laboratory’s Whitman Center, published their results in the journal Optica.

“Everybody knows fluorescence imaging is inefficient in that the microscope only captures a portion of the light (spewing off the specimen),” said senior author Hari Shroff of the National Institute of Biomedical Imaging and Bioengineering. “In this paper, we showed you can not only capture that lost light, but use computation to fuse it to the existing image and make the image sharper.”

Developed by Yicong Wu, a staff scientist in Shroff’s lab, the new system achieved resolution of up to 235 x 235 x 340 nanometers—double the volumetric resolution of traditional fluorescence microscopy methods.

To collect more of the available light, which in turn provides more information about the specimen, the new microscope has three objective lenses acquiring views of the sample simultaneously. The views are then aligned and merged by a computational process known as deconvolution.

Those computations were worked out in collaboration with co-author Patrick La Rivière of the University of Chicago’s Department of Radiology, who typically develops algorithms for improving “dose efficiency” in human-scale medical imaging, such as CAT scans.

“In medical imaging, we are always worried about dose, about capturing every X-ray. We are concerned with ‘How can we do more with less?’” La Rivière said.

In microscopy, the amount of light used presents similar concerns. “If you use very intense illuminations to image something microscopic like a worm embryo, you might change its biology or even kill it. You need to be dose efficient with your light,” La Rivière said.

La Rivière and Shroff began collaborating at the MBL in 2014, initially on algorithms to improve Shroff’s dual-view microscope, which has two objective lenses, and eventually on the new triple-view, three-lensed microscope.

La Rivière was named an MBL Fellow this year. Shroff is a Whitman Center Scientist and co-director of the MBL’s Optical Microscopy and Imaging in the Biomedical Sciences course.

Citation: Yicong Wu, P. Chandris, P.W. Winter, E.Y. Kim, V. Jaumouillé, A. Kumar, M. Guo, J.M. Leung, C. Smith, I. Rey-Suarez, H. Liu, C.M. Waterman, K.S. Ramamurthi, P. La Riviere, H. Shroff (2016) Simultaneous multi-view capture and fusion improves spatial resolution in wide-field and light-sheet microscopy, Optica, Vol. 3, Issue 8, pp. 897-910 (2016) , DOI: 10.1364/OPTICA.3.000897