Multiscale fluorescence imaging of living samples

Wu, Yicong; Shroff, Hari

doi:10.1007/s00418-022-02147-4

Cited by 8 publications

(5 citation statements)

References 101 publications

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“…The window we analyze contains relatively fast, microscale dynamics that would correspond to bacterial motility, cell signaling, and protein expression. By limiting our analysis to this window, we can also exclude slower plant processes such as osmotic transport, plant exudation, energy-storage granule formation, and plant cell division 43 . Further, while we focus our analysis here on a 0.1–4 Hz sampling speed, future experiments may opt for faster or slower imaging speeds to capture different dynamic processes.…”

Section: Discussionmentioning

confidence: 99%

Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy

Filan,

Green,

Diering

et al. 2024

Sci Rep

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The increasing global demand for food, coupled with concerns about the environmental impact of synthetic fertilizers, underscores the urgency of developing sustainable agricultural practices. Nitrogen-fixing bacteria, known as diazotrophs, offer a potential solution by converting atmospheric nitrogen into bioavailable forms, reducing the reliance on synthetic fertilizers. However, a deeper understanding of their interactions with plants and other microbes is needed. In this study, we introduce a recently developed label-free 3D quantitative phase imaging technology called dynamic quantitative oblique back-illumination microscopy (DqOBM) to assess the functional dynamic activity of diazotrophs in vitro and in situ. Our experiments involved three different diazotrophs (Sinorhizobium meliloti, Azotobacter vinelandii, and Rahnella aquatilis) cultured on media with amendments of carbon and nitrogen sources. Over 5 days, we observed increased dynamics in nutrient-amended media. These results suggest that the observed bacterial dynamics correlate with their metabolic activity. Furthermore, we applied qOBM to visualize microbial dynamics within the root cap and elongation zone of Arabidopsis thaliana primary roots. This allowed us to identify distinct areas of microbial infiltration in plant roots without the need for fluorescent markers. Our findings demonstrate that DqOBM can effectively characterize microbial dynamics and provide insights into plant-microbe interactions in situ, offering a valuable tool for advancing our understanding of sustainable agriculture.

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Section: Discussionmentioning

confidence: 99%

Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy

Filan,

Green,

Diering

et al. 2024

Sci Rep

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“…The modern technologies used in microscopic research in biology make it possible to address a wide range of problems: from monitoring of the development of the whole embryos, through the localization of single molecules in a cell, to direct visualization of the structure of macromolecules in their native state [ 1 , 2 , 3 ]. The development of both light and electron microscopy is constantly expanding the range of possibilities for researchers; however, despite all the successes achieved so far, both approaches retain their fundamental limitations.…”

Section: Principles Of Sxm and How It Compares With Other Types Of Mi...mentioning

confidence: 99%

Soft X-ray Microscopy in Cell Biology: Current Status, Contributions and Prospects

Golyshev,

Kazakov,

Kireev

et al. 2024

Acta Naturae

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The recent advances achieved in microscopy technology have led to a significant breakthrough in biological research. Super-resolution fluorescent microscopy now allows us to visualize subcellular structures down to the pin-pointing of the single molecules in them, while modern electron microscopy has opened new possibilities in the study of protein complexes in their native, intracellular environment at near-atomic resolution. Nonetheless, both fluorescent and electron microscopy have remained beset by their principal shortcomings: the reliance on labeling procedures and severe sample volume limitations, respectively. Soft X-ray microscopy is a candidate method that can compensate for the shortcomings of both technologies by making possible observation of the entirety of the cellular interior without chemical fixation and labeling with an isotropic resolution of 40–70 nm. This will thus bridge the resolution gap between light and electron microscopy (although this gap is being narrowed, it still exists) and resolve the issue of compatibility with the former, and possibly in the near future, the latter methods. This review aims to assess the current state of soft X-ray microscopy and its impact on our understanding of the subcellular organization. It also attempts to look into the future of X-ray microscopy, particularly as relates to its seamless integration into the cell biology toolkit.

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“…Professor Mühlfeld’s award manuscript was published in 2021 (Mühlfeld 2021 ), and we now have the honor to present Professor Shrof’s prize-winning award manuscript. In this beautifully crafted and illustrated review, together with Yicong Wu, Professor Shrof presents a state-of-the-art synopsis of multiscale (representing temporal and spatial scales) fluorescence imaging of living samples, ranging from cells to organisms to animals (Wu and Shrof 2022 ). The review is divided into sections describing (1) overall challenges and basic considerations in imaging fluorescent tags in living samples; (2) an “instrument” section in which the myriad of available technologies are concisely explained through examples from the literature illustrating their successful application in multiscale fluorescence imaging; and (3) suggestions and tips for those planning their own multiscale fluorescence imaging of living samples.…”

Section: Life In Technicolormentioning

confidence: 99%