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Improving microscopy by following the astronomers' guide star
Date:2/17/2011

clearer view of the target. Today, adaptive optics instrumentation accompanies all of the world's major telescopes.

Unlike in astronomy, microscopists can't place a wavefront sensor within a live animal to directly measure the distortions of light deep within tissue. To get around this problem, Betzig reasoned that the perfect focus is nothing more than a bunch of rays converging from many different directions to the same identical point. The heterogeneity of tissue means that ray is deflected differently so they no longer meet at a single point. Betzig figured that if they could study the rays individually, they could correct their deflections and steer them back to a single focus.

For their 2009 study, Betzig and Ji buried fluorescent beads underneath thick slices of mouse brain. The beads act as guide stars to help measure the deflections of the rays. To do that, the pair use a one-inch display called a spatial light modulator. The display allows them to turn on one ray at a time and then take an image of the bead. They can then determine how much the ray is deflected from the amount the bead's image is shifted relative to the desired focal point. The display is then used like a small, tiltable mirror to steer the ray back to the focal point. The process is then repeated with each of the other rays. This strategy improves the fluorescent signal and recovers optimal resolution through a chunk of tissue up to 400 micrometers thick. "Another advantage is that it's very efficient in terms of how little light is needed," Betzig says. "Light is not completely noninvasive, so as microscopists we have to be very careful not to damage our specimens."

Ji, who will soon begin her own lab as a fellow at Janelia Farm, has recently taken the technique to the next level: live brain imaging in mice. To do this, she uses genetic engineering to label the brains of mouse fetuses with a fluorescent marker of neurons while at the same time injecting
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Contact: Andrea Widener
widenera@hhmi.org
301-215-8807
Howard Hughes Medical Institute
Source:Eurekalert

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