Joint-tagging Super-resolution Optical Fluctuation Imaging: Live-cell imaging enables super resolution in space and time

Super-resolution microscopy methods provide sub-diffractional spatial resolution, and thereby enable visualization of subcellular organelles.

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Super-resolution microscopy methods provide sub-diffractional spatial resolution, and thereby enable visualization of subcellular organelles. But high spatial resolution is typically achieved at the expense of temporal resolution. And the methods generally involve complicated and costly instrumentation, which hinders their wide application.

Now, a research team at Peking University (Beijing, China) has demonstrated a novel technique designed to provide high spatial and temporal resolution-simultaneously. Using light's spectral characteristic, the researchers have combined three techniques: Quantum dot (QD), spectrum isolation, and super-resolution optical fluctuation imaging (SOFI).

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Joint tagging involves low densities of multiple types of quantum dots (a) compared to single tagging, which involves ultra-high densities of a single type (b). SOFI processing completes the operation.

The method is simple, according to Peng Xi, Ph.D., Associate Professor in the Department of Biomedical Engineering: In a technique called "joint tagging" (JT), the subcelluar organelle is labeled with multiple types of QDs—and then imaged using separate color channels (simultaneous excitation is possible because of the unique blue-shifted absorption spectrum of QDs). The QDs' fluorescence emission is characterized by blinking and by a narrow spectrum. The dots can be detected by individual color channel and analyzed with a single-molecule statistical-based super-resolution technique called super-resolution optical fluctuation imaging (SOFI).

The approach enables spatiotemporal resolution up to 3 s and 85 nm using a conventional, wide-field fluorescence microscope.1 Joint tagging with multiple QDs at one subcellular target enables suppression of artefacts, and results in high-fidelity image reconstruction. Further, as the spectrally isolated QDs are closer to single-molecule status,2 JT-SOFI can largely improve the temporal resolution of super-resolution imaging. It therefore provides a novel solution for imaging live cells.

1. Z. Zeng et al., Sci. Rep., 5, 8359 (2015); doi:10.1038/srep08359.

2. E. Betzig, Opt. Lett., 20, 3, 237–239 (1995).

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