Label-free fluorescence microscopy in fungi

Knaus, Helene; Blab, Gerhard A.; Veluw, G. Jerre van; Gerritsen, Hans C.; Wösten, Han A. B.

doi:10.1016/j.fbr.2013.05.003

Cited by 26 publications

(19 citation statements)

References 76 publications

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“…Microscopy allows to gain new insights at high spatial and temporal resolution into essential cellular processes such as protein localization, physiological activity, and growth dynamics (Hickey et al, 2004;Knaus et al, 2013;Chapuis et al, 2019). However, fluorescence microscopy of fungi is limited by the small size of their organelles, which is below the diffraction-limited resolution provided by conventional fluorescence microscopes.…”

Section: Introductionmentioning

confidence: 99%

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Expansion Microscopy for Cell Biology Analysis in Fungi

Götz¹,

Panzer²,

Trinks³

et al. 2020

Front. Microbiol.

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Super-resolution microscopy has evolved as a powerful method for subdiffractionresolution fluorescence imaging of cells and cellular organelles, but requires sophisticated and expensive installations. Expansion microscopy (ExM), which is based on the physical expansion of the cellular structure of interest, provides a cheap alternative to bypass the diffraction limit and enable super-resolution imaging on a conventional fluorescence microscope. While ExM has shown impressive results for the magnified visualization of proteins and RNAs in cells and tissues, it has not yet been applied in fungi, mainly due to their complex cell wall. Here we developed a method that enables reliable isotropic expansion of ascomycetes and basidiomycetes upon treatment with cell wall degrading enzymes. Confocal laser scanning microscopy (CLSM) and structured illumination microscopy (SIM) images of 4.5-fold expanded sporidia of Ustilago maydis expressing fluorescent fungal rhodopsins and hyphae of Fusarium oxysporum or Aspergillus fumigatus expressing either histone H1-mCherry together with Lifeact-sGFP or mRFP targeted to mitochondria, revealed details of subcellular structures with an estimated spatial resolution of around 30 nm. ExM is thus well suited for cell biology studies in fungi on conventional fluorescence microscopes.

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Section: Introductionmentioning

confidence: 99%

“…However, fluorescence microscopy of fungi is limited by the small size of their organelles, which is below the diffraction-limited resolution provided by conventional fluorescence microscopes. In addition, fungi tend to exhibit strong autofluorescence (Knaus et al, 2013), further complicating high-end fluorescence imaging.…”

Section: Introductionmentioning

confidence: 99%

Expansion Microscopy for Cell Biology Analysis in Fungi

Götz¹,

Panzer²,

Trinks³

et al. 2020

Front. Microbiol.

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show abstract

“…Furthermore, the CLSM images, which reveal an uneven and complex surface topology, is of limited use in understanding the 3D distribution of the spores and hyphae embedded deep in the growth medium. To overcome these drawbacks, we employed label‐free light sheet fluorescence microscopy (LSFM) for imaging the intact fungal culture . This mode of imaging is preferred, as any possible toxicity due to exogenous labeling is avoided while also precluding the need to selectively induce fluorescent protein expression within the fungi.…”

Section: Resultsmentioning

confidence: 99%

Exploring Morphological and Biochemical Linkages in Fungal Growth with Label‐Free Light Sheet Microscopy and Raman Spectroscopy

et al. 2016

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Imaging tip growth in fungal hyphae is highly warranted to unravel the molecular mechanism of this extraordinarily precise and localized phenomenon. In situ probing of fungal cultures, however, have been challenging due to their inherent complexity and light penetration issues associated with conventional optical imaging. In this work, we report a label-free approach using a combination of light sheet microscopy and Raman spectroscopy to obtain concomitant morphological and biochemical information from the growing specimen. We show that the variance in morphology in the symbiotic fungus Piriformospora indica are rooted in the underlying differences in chemical composition in the specific growth zones. Our findings suggest that this potent two-pronged approach can comprehensively characterize growth areas and elucidate microbe interactions in still developing colonies with high sensitivity and multiplexing capability.

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“…Fluorescence spectroscopy [ 209,213 ] most often measures light emission from samples with electrons that have been excited, however, it is also possible to measure absorption for cases involving single/pairs of fluorophores. [ 214 ] Fluorescence spectroscopy has been used for a variety of fundamental biochemistry studies, [ 132,215 ] for example, fluorescent dye binding assays were used to study functional amyloids that form filaments promoting eumelanin deposition (and thereby pigmentation in mammals); [ 152,216–218 ] analysis of metabolic activity of fungi (e.g., DHN–melanin and pyomelanin production by Aspergillus niger and Agaricus bisporus ), [ 219–221 ] and autofluorescence of melanins from cuttlefish ( Sepia officinalis ) and black human hair [ 222 ] and other sources. [ 223–226 ]…”

Section: Analysis Of Melaninsmentioning

confidence: 99%

Melanins as Sustainable Resources for Advanced Biotechnological Applications

et al. 2020

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Melanins are a class of biopolymers that are widespread in nature and have diverse origins, chemical compositions, and functions. Their chemical, electrical, optical, and paramagnetic properties offer opportunities for applications in materials science, particularly for medical and technical uses. This review focuses on the application of analytical techniques to study melanins in multidisciplinary contexts with a view to their use as sustainable resources for advanced biotechnological applications, and how these may facilitate the achievement of the United Nations Sustainable Development Goals.

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Label-free fluorescence microscopy in fungi

Cited by 26 publications

References 76 publications

Expansion Microscopy for Cell Biology Analysis in Fungi

Expansion Microscopy for Cell Biology Analysis in Fungi

Exploring Morphological and Biochemical Linkages in Fungal Growth with Label‐Free Light Sheet Microscopy and Raman Spectroscopy

Melanins as Sustainable Resources for Advanced Biotechnological Applications

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