Physicists at Stanford have developed a technique called multi-pass microscopy to obtain clearer images while reducing the damage inflicted on the sample. They outline their new technique in a paper titled “Multi-pass microscopy,” recently published in “Nature Communications.”

Normal light microscopes have light pass through the sample only once. The Mark Kasevich Research Group uses mirrors to reflect light back and forth across the specimen before finally being collected at a fixed number of passes.

By passing the same photons of light multiple times across the specimen, these researchers are able to improve the signal to noise ratio of the resulting image. This improves the quality of the image without using more light, which would damage the specimens.

“We try to get as much information as we can with as few interactions as we need to do,” said Thomas Juffmann, a postdoctoral research fellow and co-author of the paper. “Because an interaction always creates damage, always takes time, and so forth.”

One disadvantage of this new technique is the complexity involved with its set-up, since the microscope requires additional mirrors in order to reflect the light back and forth.

“We need basically twice as much equipment,” Juffmann said. “But what’s worse than that is that the alignment gets much harder because we need to make sure a spot on the sample… is re-imaged to exactly that spot many many times.”

Currently, the greatest difficulty with the technique is finding a way to collect the light at the end.

“How do you get the light out of your microscope because you have to have a mirror on each end so the light is kind of stuck in there?” said co-author Brannon Klopfer, a graduate student in the Kasevich group. “The easiest way of doing that is to have… a partially reflective mirror, and then you just collect this leaked out light.”

This both causes light to leak during the multi-pass process and reduces the amount of light that is used to create the final image. The Kasevich group hopes to develop a technique that reduces the light wasted in order to increase the amount captured in the end.

They also hope to eventually expand their research to electron microscopes, where the potential of damage poses a greater problem.

“You use electron microscopy when you wish to look at really small things, smaller than the wavelength of light, and it works great,” Juffmann said. “You can see single atoms in it, but you cannot image DNA or cells or proteins [at atomic resolution] simply because you create too much damage. You basically [destroy] your sample before you get a good image, and that’s where our scheme could help.”

This research takes advantage of camera technology with extremely fast shutter speeds: Shutter speeds can be as fast as 0.1 billionths of a second. This allows researchers to capture the light after it has made a fixed number of passes through the specimen.

Juffmann also uses this camera technology for an art and science outreach project called SEEC, which creates videos of light spreading at the speed of light across plants, people and objects.

Contact Aulden Foltz at afoltz ‘at’ stanford.edu.