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A Strategy to Lengthen the On-time of Photochromic Rhodamine Spirolactam for Super-Resolution Photoactivated Localization Microscopy.

08:00 EDT 2nd April 2019 | BioPortfolio

Summary of "A Strategy to Lengthen the On-time of Photochromic Rhodamine Spirolactam for Super-Resolution Photoactivated Localization Microscopy."

Rhodamine derivatives and analogues have been widely used for different super-resolution imaging techniques, including photoactivated localization microscopy (PALM). Among them, rhodamine spirolactams exhibit great superiority for PALM imaging due to a desirable bright-dark contrast during the photochromic switching process. Although considerable attention has been paid to the chemical modifications on rhodamine spirolactams, the on-time of photochromic switching, one of the key characteristics for PALM imaging, has never been optimized in previous developments. In this study, we proposed that simply installing a carboxyl group close to the lactam site could impose an intramolecular acidic environment, stabilize the photoactivated zwitterionic structure and thus effectively increase the on-time. Based on this idea, we have synthesized a new rhodamine spirolactam, Rh-Gly, that demonstrated considerably longer on-time than the other tested analogues, as well as an enhancement of single molecule brightness, an improvement on signal-to-noise ratio and an enlargement of total collected photons of single molecule before photobleach. Finally, super-resolution images of live cell mitochondria stained with Rh-Gly have been obtained with a good temporal resolution of 10 s, as well as a satisfactory localization precision of ~25 nm. Through self-labeling protein tags, Rh-Gly modified with HaloTag ligand enabled super-resolution imaging of histone H2B proteins in live HeLa cells; through immunostaining antibodies labeled with an isothiocyanate-substituted Rh-Gly, super-resolution imaging of microtubules was achieved in fixed cells. Therefore, our simple and effective strategy provides novel insight for developing further enhanced rhodamine spirolactams recommendable for PALM imaging.

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This article was published in the following journal.

Name: Journal of the American Chemical Society
ISSN: 1520-5126
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