Photoactivatable fluorescent proteins for super-resolution microscopy

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Abstract

Super-resolution fluorescence microscopy techniques such as simulated emission depletion (STED) microscopy and photoactivated localization microscopy (PALM) allow substructures, organelles or even proteins within a cell to be imaged with a resolution far below the diffraction limit of ~200 nm. The development of advanced fluorescent proteins, especially photoactivatable fluorescent proteins of the GFP family, has greatly contributed to the successful application of these techniques to live-cell imaging. Here, we will illustrate how two fluorescent proteins with different photoactivation mechanisms can be utilized in high resolution dual color PALM imaging to obtain insights into a cellular process that otherwise would not be accessible. We will explain how to set up and perform the experiment and how to use our latest software “a-livePALM” for fast and efficient data analysis.

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Ishitsuka, Y., Nienhaus, K., & Nienhaus, G. U. (2014). Photoactivatable fluorescent proteins for super-resolution microscopy. Methods in Molecular Biology, 1148, 239–260. https://doi.org/10.1007/978-1-4939-0470-9_16

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