MICROSCOPY/NEUROIMAGING: Third-harmonic generation enables label-free imaging of live brain tissue

The ability to visualize neurons inside living brain tissue is a fundamental requirement in neuroscience and neurosurgery-and a noninvasive probe capable of micrometer-scale resolution is on the wish list of every neurologist.

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The ability to visualize neurons inside living brain tissue is a fundamental requirement in neuroscience and neurosurgery—and a noninvasive probe capable of micrometer-scale resolution is on the wish list of every neurologist. While two-photon laser-scanning microscopy (2PLSM) has become the standard for minimally invasive high-resolution imaging of living biological samples, the approach requires the use of fluorescent dyes to provide image contrast—which is less than ideal. Now, a team of Dutch scientists has demonstrated high-contrast imaging of live brain tissue at cellular resolution, without the need for fluorescent probes, using optical third-harmonic generation (THG).

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How the technique works: By changing the focus of the laser beam, researchers can visualize neuronal projections without fluorescent dyes.

The researchers say they exploited the specific geometry and lipid content of brain tissue at the cellular level to achieve partial phase matching of THG, providing an alternative contrast mechanism to fluorescence. “We find that THG brain imaging allows rapid, noninvasive label-free imaging of neurons, white-matter structures, and blood vessels simultaneously,” they report. “Furthermore, we exploit THG-based imaging to guide micropipettes towards designated neurons inside live tissue.”

This work represents a step toward label-free microscopic live brain imaging, and opens up possibilities for the development of laser-guided microsurgery techniques in the living brain. And because abnormal lipid metabolism characterizes Alzheimer’s disease and brain cancer, the tool could detect these medical conditions.

1. S. Witte et al., PNAS 108 (15), 5970–5975 (2011).

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