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Ideal fluorophores for STORM should be very bright, have a high rate of photoswitching and exhibit minimal photobleaching in thiol-containing buffers. The first one, known as direct STORM or dSTORM microscopy, is compatible with many commonly used fluorophores, which can be converted to an off state using specific excitation parameters and in combination with specialized oxygen-scavenging imaging buffers. What kind of fluorophores can be used for STORM microscopy? For a successful STORM imaging, individual fluorophores must be sparse enough, so that only one molecule is activated within a diffraction-limited region at any given time.ĭuring the STORM reconstruction process, individual molecules can be precisely localized by determining their position coordinates from the photons detected for each activation event (often as a Gaussian function). This process is sequentially repeated many times until most fluorophores have been imaged.
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STORM stands for “Stochastic Optical Reconstruction Microscopy” and it relies on the stochastic activation of individual fluorophores with photoactivatable properties.ĭuring STORM, single fluorophores “blink” by a process of random activation from an off or dark state, to an on or emission state, quickly followed by a switch back to a dark state or photobleaching. Since its invention in 2006, STORM microscopy has become the most widely used super-resolution microscopy technique for single-molecule imaging.