Taking the problem of diffraction out of single-molecule localisation microscopy
Look down at your feet in the sea and they may not be where you expect – a trick of the light’s rays as they bend, or diffract at the boundary from water to air. Microscopists face a similar problem when peering into cells, and often use super-resolution fluorescence microscopy to unravel some of this confusing diffraction. But in thicker tissue samples, layer upon layer of light-bending structures leave images blurry, obscuring important detail. Yet the tiny dendrite branches of nerve cells inside this chunk of a mouse’s visual cortex are revealed clearly in 3D. The delicate structures were peppered with harmless fluorescent molecules which blink like tiny beacons, revealing their 3D coordinates through the haze to high-powered microscopes. Using computer software to join the dots and recreate the nanoscale scene, these techniques could be used routinely with thicker tissue samples like biopsies to reveal clues about neurodegenerative diseases or cancer.
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