Live‐cell RESOLFT nanoscopy of transgenic <i>Arabidopsis thaliana</i>

Schnorrenberg, Sebastian; Ghareeb, Hassan; Frahm, Lars; Grotjohann, Tim; Jensen, Nickels; Teichmann, Thomas; Hell, Stefan W.; Lipka, Volker; Jakobs, Stefan

doi:10.1002/pld3.261

Cited by 8 publications

(8 citation statements)

References 38 publications

(58 reference statements)

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“…Critically, prominent cell walls and unique organelles like vacuoles produce refractive index variations and pronounced light scattering effects 3 . However, the plant community has started to overcome these challenges to advance our understanding of subcellular plant biology 4,5 , with application of techniques like structured illumination microscopy (SIM) 6–11 , stimulated emission depletion (STED) 12,13 and REversible Saturable Optical Linear Fluorescence Transitions (RESOLFT) 14 microscopy, as well as single molecule based methods including photo-activated localization microscopy (PALM) 8,15,16 and stochastic optical reconstruction microscopy (STORM) 17,18 . As much as these technologies have advanced our ability to visualize plants at improved resolution, some of these provide only an extension of resolution by a factor of two (linear SIM) or below (Airyscan) and hence do not access the sub-100 nm resolution range.…”

Section: Mainmentioning

confidence: 99%

Super-resolution expansion microscopy in plant roots

Gallei,

Truckenbrodt,

Kreuzinger

et al. 2024

Preprint

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Super-resolution methods enable spatial resolution far better than the optical diffraction limit of about half the wavelength of light (∼200-300 nm) but have yet to attain widespread use in plants, owing in large part to plants’ challenging optical properties. Expansion microscopy improves effective resolution by isotropically increasing physical distances between sample structures while preserving relative spatial arrangements, and clears the sample. However, its application to plants has been hindered by the rigid, mechanically cohesive structure of plant tissues. Here, we report on whole-mount expansion microscopy ofArabidopsis thalianaroot tissues (PlantEx), achieving 4-fold resolution increase over conventional microscopy, highlighting microtubule cytoskeleton organization and interaction between molecularly defined cellular constituents. By combining PlantEx with STED microscopy, we increase nanoscale resolution further and visualize the complex organization of subcellular organelles from intact tissues by example of the densely packed COPI-coated vesicles associated with the Golgi apparatus and put these into cellular structural context.

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Section: Mainmentioning

confidence: 99%

Super-resolution expansion microscopy in plant roots

Gallei,

Truckenbrodt,

Kreuzinger

et al. 2024

Preprint

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“…However, it requires the use of photo-switchable fluorophores that reversibly transit between on and off states according to illumination intensity rendering this method unsuitable for use with conventional fluorescent proteins. Nevertheless, RESOLFT being a scanning method such as STED can be used for live imaging and was recently applied in plants to visualize MTs and actin filaments ( Schnorrenberg et al, 2020 ).…”

Section: Super-resolution Microscopymentioning

confidence: 99%

Imaging plant cells and organs with light-sheet and super-resolution microscopy

et al. 2021

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The documentation of plant growth and development requires integrative and scalable approaches to investigate and spatiotemporally resolve various dynamic processes at different levels of plant body organization. The present update deals with vigorous developments in mesoscopy, microscopy and nanoscopy methods that have been translated to imaging of plant subcellular compartments, cells, tissues and organs over the past 3 years with the aim to report recent applications and reasonable expectations from current light-sheet fluorescence microscopy (LSFM) and super-resolution microscopy (SRM) modalities. Moreover, the shortcomings and limitations of existing LSFM and SRM are discussed, particularly for their ability to accommodate plant samples and regarding their documentation potential considering spherical aberrations or temporal restrictions prohibiting the dynamic recording of fast cellular processes at the three dimensions. For a more comprehensive description, advances in living or fixed sample preparation methods are also included, supported by an overview of developments in labeling strategies successfully applied in plants. These strategies are practically documented by current applications employing model plant Arabidopsis thaliana (L.) Heynh., but also robust crop species such as Medicago sativa L. and Hordeum vulgare L. Over the past few years, the trend towards designing of integrative microscopic modalities has become apparent and it is expected that in the near future LSFM and SRM will be bridged to achieve broader multiscale plant imaging with a single platform.

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“…The resolution is improved by quenching of the fluorescence at the edge of the illuminated area, with fluorescence occurring only in the unquenched area within the annulus. In addition to STED, many other different high-resolution microscopy techniques have been developed, such as reversible saturable optically linear fluorescence transitions (RESOLFTs) [2], structured illumination microscopy (SIM) [3], stochastic optical reconstruction microscopy (STORM) [4], photoactivated localization microscopy (PALM) [5], and fluorescence photoactivation localization microscopy (FPALM) [6].…”

Section: Introductionmentioning

confidence: 99%

“…1 Two or three fingers would be used to hide or make visible the second (G) or third channel (B). 2 Actions are active again for the whole objects-for all visible channels. 3 Button-pressing means that the further applied functions were later focused on a specific channel so that, for example, if button 3 is pressed, the following actions are applied to the third channel (B).…”

mentioning

confidence: 99%

A Novel Gesture-Based Control System for Fluorescence Volumetric Data in Virtual Reality

Čmiel

Chmelikova

Zumberg

et al. 2021

Sensors

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With the development of light microscopy, it is becoming increasingly easy to obtain detailed multicolor fluorescence volumetric data. The need for their appropriate visualization has become an integral part of fluorescence imaging. Virtual reality (VR) technology provides a new way of visualizing multidimensional image data or models so that the entire 3D structure can be intuitively observed, together with different object features or details on or within the object. With the need for imaging advanced volumetric data, demands for the control of virtual object properties are increasing; this happens especially for multicolor objects obtained by fluorescent microscopy. Existing solutions with universal VR controllers or software-based controllers with the need to define sufficient space for the user to manipulate data in VR are not usable in many practical applications. Therefore, we developed a custom gesture-based VR control system with a custom controller connected to the FluoRender visualization environment. A multitouch sensor disk was used for this purpose. Our control system may be a good choice for easier and more comfortable manipulation of virtual objects and their properties, especially using confocal microscopy, which is the most widely used technique for acquiring volumetric fluorescence data so far.

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Live‐cell RESOLFT nanoscopy of transgenic Arabidopsis thaliana

Abstract: This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

Cited by 8 publications

References 38 publications

Super-resolution expansion microscopy in plant roots

Super-resolution expansion microscopy in plant roots

Imaging plant cells and organs with light-sheet and super-resolution microscopy

A Novel Gesture-Based Control System for Fluorescence Volumetric Data in Virtual Reality

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