Vital Autofluorescence: Application to the Study of Plant Living Cells

Roshchina, Victoria V.

doi:10.1155/2012/124672

Cited by 79 publications

(63 citation statements)

References 51 publications

(112 reference statements)

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“…Most substances contributing to autofluorescence background in plants share an excitation maximum in the violet/blue range, whereas the emission maxima are distinct between the fluorophores and, thus, affect spectral variants of fluorescent proteins to different extents (Roshchina, 2012; Table 2). Between these substances are secondary metabolites that accumulate in the vacuole, but also ubiquitously distributed molecules such as flavins.…”

Section: Fluorescent Proteins For Fretmentioning

confidence: 99%

Quantification of Förster resonance energy transfer by monitoring sensitized emission in living plant cells

et al. 2013

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Förster resonance energy transfer (FRET) describes excitation energy exchange between two adjacent molecules typically in distances ranging from 2 to 10 nm. The process depends on dipole-dipole coupling of the molecules and its probability of occurrence cannot be proven directly. Mostly, fluorescence is employed for quantification as it represents a concurring process of relaxation of the excited singlet state S1 so that the probability of fluorescence decreases as the probability of FRET increases. This reflects closer proximity of the molecules or an orientation of donor and acceptor transition dipoles that facilitates FRET. Monitoring sensitized emission by 3-Filter-FRET allows for fast image acquisition and is suitable for quantifying FRET in dynamic systems such as living cells. In recent years, several calibration protocols were established to overcome to previous difficulties in measuring FRET-efficiencies. Thus, we can now obtain by 3-filter FRET FRET-efficiencies that are comparable to results from sophisticated fluorescence lifetime measurements. With the discovery of fluorescent proteins and their improvement toward spectral variants and usability in plant cells, the tool box for in vivo FRET-analyses in plant cells was provided and FRET became applicable for the in vivo detection of protein-protein interactions and for monitoring conformational dynamics. The latter opened the door toward a multitude of FRET-sensors such as the widely applied Ca2+-sensor Cameleon. Recently, FRET-couples of two fluorescent proteins were supplemented by additional fluorescent proteins toward FRET-cascades in order to monitor more complex arrangements. Novel FRET-couples involving switchable fluorescent proteins promise to increase the utility of FRET through combination with photoactivation-based super-resolution microscopy.

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Section: Fluorescent Proteins For Fretmentioning

confidence: 99%

Quantification of Förster resonance energy transfer by monitoring sensitized emission in living plant cells

et al. 2013

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“…These pteridine ring-containing compounds would likely contribute to the fluorescence we observe in the FITC channels, but are by no means the only candidates for the autofluorescence that we observe. Candidate natural autofluorophores have been reviewed comprehensively (37). Universal cellular fluorophores in the blue-green region of the spectrum are NADP(H), pterins, and flavins (38,39).…”

Section: Discussionmentioning

confidence: 99%

Fluorescence Imaging of Posterior Spiracles from Second and Third Instars of Forensically Important Chrysomya rufifacies (Diptera: Calliphoridae)^,

Flores

Miller

Showman

et al. 2016

Journal of Forensic Sciences

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Entomological protocols for aging blow fly (Diptera: Calliphoridae) larvae to estimate the time of colonization (TOC) are commonly used to assist in death investigations. While the methodologies for analysing fly larvae differ, most rely on light microscopy, genetic analysis or, more rarely, electron microscopy. This pilot study sought to improve resolution of larval stage in the forensically-important blow fly Chrysomya rufifacies using high-content fluorescence microscopy and biochemical measures of developmental marker proteins. We established fixation and mounting protocols, defined a set of measurable morphometric criteria and captured developmental transitions of 2nd instar to 3rd instar using both fluorescence microscopy and anti-ecdysone receptor Western blot analysis. The data show that these instars can be distinguished on the basis of robust, non-bleaching, autofluorescence of larval posterior spiracles. High content imaging techniques using confocal microscopy, combined with morphometric and biochemical techniques, may therefore aid forensic entomologists in estimating TOC.

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“…It is also noteworthy that the emission pattern of a compound in an in situ situation may exhibit some distinctions in comparison to an in vitro assay, since the latter is obtained by purified standard, while the target compound is stabilized by different ligands in the cell compartments (Roshchina, 2008(Roshchina, , 2012.…”

Section: Considerations On the Polyphenol Analysis By Confocal Microsmentioning

confidence: 99%

Synergism between ozone and light stress: structural responses of polyphenols in a woody Brazilian species

2016

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Microscopic studies on isolated ozone (O3) effects or on those in synergy with light stress commonly report the induction of polyphenols that exhibit different aspects within the vacuole of photosynthesizing cells. It has been assumed that these different aspects are randomly spread in the symptomatic (injured) regions of the leaf blade. Interestingly, secretory ducts that constitutively produce polyphenols also exhibit these same variations in their vacuolar aspect, in a spatial sequence related to the destiny of these cells (e.g., programmed cell death (PCD) in lytic secretion processes). Here, we demonstrate that the deposition pattern of polyphenols prior to the establishment of the hypersensitive-like response, a type of PCD caused by O3, follows the same one observed in the epithelial cells of the constitutive lysigenous secretory ducts. Astronium graveolens, an early secondary Brazilian woody species, was selected based on its susceptibility to high light and presence of secretory ducts. The synergism effects were assessed by exposing plants to the high O3 concentrations at an urban site in São Paulo City. Confocal, widefield and light microscopies were used to examine polyphenols' occurrence and aspects. The spatial pattern of polyphenols distribution along the leaflets of plants submitted to the synergism condition, in which a dense vacuolar aspect is the target of a cell destined to death, was also observed in the constitutive secretory cells prior to lysis. This similar structural pattern may be a case of homology of process involving both the constitutive (secretory ducts) and the induced (photosynthesizing cells) defenses.

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Vital Autofluorescence: Application to the Study of Plant Living Cells

Cited by 79 publications

References 51 publications

Quantification of Förster resonance energy transfer by monitoring sensitized emission in living plant cells

Quantification of Förster resonance energy transfer by monitoring sensitized emission in living plant cells

Fluorescence Imaging of Posterior Spiracles from Second and Third Instars of Forensically Important Chrysomya rufifacies (Diptera: Calliphoridae)^,

Synergism between ozone and light stress: structural responses of polyphenols in a woody Brazilian species

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Vital Autofluorescence: Application to the Study of Plant Living Cells

Cited by 79 publications

References 51 publications

Quantification of Förster resonance energy transfer by monitoring sensitized emission in living plant cells

Quantification of Förster resonance energy transfer by monitoring sensitized emission in living plant cells

Fluorescence Imaging of Posterior Spiracles from Second and Third Instars of Forensically Important Chrysomya rufifacies (Diptera: Calliphoridae),

Synergism between ozone and light stress: structural responses of polyphenols in a woody Brazilian species

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Fluorescence Imaging of Posterior Spiracles from Second and Third Instars of Forensically Important Chrysomya rufifacies (Diptera: Calliphoridae)^,