Quantum Dot Blueing and Blinking Enables Fluorescence Nanoscopy

Hoyer, Patrick; Staudt, Thorsten; Engelhardt, Johann; Hell, Stefan W.

doi:10.1021/nl103639f

Cited by 101 publications

(106 citation statements)

References 35 publications

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“…These go back to Lidke et al, whose work on independent component analysis with quantum dots presaged STORM/PALM type localization-based microscopy [122]. Subsequent efforts have found application of quantum dots to imaging by RESOLFT, STORM/PALM, and SOFI although they have not found as frequent use as fluorescent proteins or organic fluorophroes [120,123,124]. One concern regarding the use of core-shell quantum dots (which are also surface-passivated and functionalized) is their relatively large hydrodynamic radius ($20 nm), although this may potentially be improved by the use of smaller quantum dots in the future [125].…”

Section: Additional Super-resolution Techniques and Fluorescent Probesmentioning

confidence: 99%

Twinkle, twinkle little star: Photoswitchable fluorophores for super‐resolution imaging

2014

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a b s t r a c tPhotoswitchable fluorescent probes are key elements of newly developed super-resolution fluorescence microscopy techniques that enable far-field interrogation of biological systems with a resolution of 50 nm or better. In contrast to most conventional fluorescence imaging techniques, the performance achievable by most super-resolution techniques is critically impacted by the photoswitching properties of the fluorophores. Here we review photoswitchable fluorophores for superresolution imaging with discussion of the fundamental principles involved, a focus on practical implementation with available tools, and an outlook on future directions.

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Section: Additional Super-resolution Techniques and Fluorescent Probesmentioning

confidence: 99%

Twinkle, twinkle little star: Photoswitchable fluorophores for super‐resolution imaging

2014

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“…Understanding the principle of these optical processes are necessary, not only toward minimizing or compensating for their effects (Uematsu et al 2006;Mahler et al 2008;Wang et al 2009), but toward controlling their synthesis (Dabbousi et al 1997;He et al 2006;Mandal and Tamai 2011), or exploiting new application (Tonti et al 2004;Antelman et al 2009;Chien et al 2011;Hoyer et al 2011;Shi et al 2012).…”

Section: Introductionmentioning

confidence: 99%

“…These reports provided a good deal of scientific insight into the effect of different environments on photoluminescence properties. For a new promising fluorescent probes, the photo-induced effects must be considered, since photo-induced QDs properties are important for QDs synthesis (Liu et al 2008;Sun et al 2014) and applications (Hoyer et al 2011;Shi et al 2014). The photo-induced effects on the fluorescence intermittency and photoluminescence enhancement have been attracting widespread concerns (Tonti et al 2004;Uematsu et al 2005;Crouch et al 2009;Zhang et al 2013;Hoy et al 2014).…”

Section: Introductionmentioning

confidence: 99%

“…Higher laser intensities accelerated the process of blue shift, meanwhile, slow spectral blue shifts were also observed under low-level illumination from a 3 W cm -2 Xe arc lamp (Cordero et al 2000). Exciting QDs by appropriate excitation light source can obtain expected rate and extent of blue shift that enables QDs to fluorescence imaging beyond the diffraction-limit (Hoyer et al 2011;Shi et al 2014). But, as shown in the above paper, they did not reveal the oxidation kinetics of single QDs and did not achieve quantitative comparison of the photo-oxidation of single QDs at different excitation wavelengths and intensities.…”

Section: Introductionmentioning

confidence: 99%

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Excitation wavelength and intensity dependence of photo-spectral blue shift in single CdSe/ZnS quantum dots

et al. 2014

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The influence of excitation wavelength and intensity on core/shell CdSe/ZnS quantum dots (QDs) photo-spectral blue shift was investigated by spectral imaging. Analysis of the evolution of the distance between the zeroth-order spot and the firstorder spectral streak, we found that the extent of blue shift strongly depends on the excitation wavelength and QDs sizes, but not on the excitation intensity. Converted the extent of blue shift into the decreased QDs volume at a series of time, the core oxidation kinetics of CdSe/ZnS QDs was uncovered that provided a quantitative comparison method for study the excitation wavelength and intensity dependence of single QDs blue shift. The core oxidation rate is almost proportional to the excitation intensity. These results are explained by a fact that higher energy excitation wavelength can accelerate individual exciton formation and higher excitation intensity can induce more amount of exciton formation per a unit time.

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“…37 for Dendra2 in comparison to organic fluorophores characterized in literature, such as Alexa488 (49,50), Alexa546 and Alexa555 (51), and QD565, a type of CdSe quantum dots (QDs) (51). Of course, to use QDs, one would need to establish a method of photoactivation/photoconversion that results from a single-photon absorption event, or some other method of controlling the fraction of QDs in a dark state (52), such that the majority are dark at a given time. The point, however, is that several particular photophysical properties of a fluorophore determine the best resolution (by a chosen definition) that can, in principle, be obtained in a localization microscope.…”

Section: Photophysics and Localization Microscopymentioning

confidence: 99%

The Role of Probe Photophysics in Localization-Based Superresolution Microscopy

Pennacchietti

Gould

Hess

2018

Biophysical Journal

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Fluorescent proteins are used extensively for biological imaging applications; photoactivatable and photoconvertible fluorescent proteins (PAFPs) are used widely in superresolution localization microscopy methods such as fluorescence photoactivation localization microscopy and photoactivated localization microscopy. However, their optimal use depends on knowledge of not only their bulk fluorescence properties, but also their photophysical properties at the single molecule level. We have used fluorescence correlation spectroscopy and cross-correlation spectroscopy to quantify the diffusion, photobleaching, fluorescence intermittency, and photoconversion dynamics of Dendra2, a well-known PAFP used in localization microscopy. Numerous dark states of Dendra2 are observed both in inactive (green fluorescent) and active (orange fluorescent) forms; the interconversion rates are both light-and pH-dependent, as observed for other PAFPs. The dark states limit the detected count rate per molecule, which is a crucial parameter for localization microscopy. We then developed, to our knowledge, a new mathematical estimate for the resolution in localization microscopy as a function of the measured photophysical parameters of the probe such as photobleaching quantum yield, count rate per molecule, and intensity of saturation. The model was used to predict the dependence of resolution on acquisition parameters such as illumination intensity and time per frame, demonstrating an optimal set of acquisition parameters for a given probe for a variety of measures of resolution. The best possible resolution was then compared for Dendra2 and other widely used probes, including Alexa dyes and quantum dots. This work establishes a framework for determination of the best possible resolution using a localization microscope to image a particular fluorophore, and suggests that development of probes for use in superresolution localization microscopy must consider the count rate per molecule, the saturation intensity, the photobleaching yield, and, crucially, management of bright/dark state transitions, to optimize image resolution.

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Quantum Dot Blueing and Blinking Enables Fluorescence Nanoscopy

Cited by 101 publications

References 35 publications

Twinkle, twinkle little star: Photoswitchable fluorophores for super‐resolution imaging

Twinkle, twinkle little star: Photoswitchable fluorophores for super‐resolution imaging

Excitation wavelength and intensity dependence of photo-spectral blue shift in single CdSe/ZnS quantum dots

The Role of Probe Photophysics in Localization-Based Superresolution Microscopy

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