scholarly journals State transitions and photosystems spatially resolved in individual cells of the cyanobacterium Synechococcus elongatus

2021 ◽  
Author(s):  
Ahmad Farhan Bhatti ◽  
Diana Kirilovsky ◽  
Herbert van Amerongen ◽  
Emilie Wientjes
Keyword(s):  
Single Cells ◽  
Synechococcus Elongatus ◽  
State Transitions ◽  
Fluorescence Lifetime Imaging ◽  
Time Resolved ◽  
Fluorescence Kinetics ◽  
Lifetime Imaging ◽  
Spatially Resolved ◽  
Acclimation Response ◽  
Kinetics Of

Abstract State transitions are a low-light acclimation response through which the excitation of Photosystem I (PSI) and Photosystem II (PSII) is balanced; however, our understanding of this process in cyanobacteria remains poor. Here, picosecond fluorescence kinetics was recorded for the cyanobacterium Synechococcus elongatus using fluorescence lifetime imaging microscopy (FLIM), both upon chlorophyll a and phycobilisome (PBS) excitation. Fluorescence kinetics of single cells obtained using FLIM were compared with those of ensembles of cells obtained with time-resolved fluorescence spectroscopy. The global distribution of PSI and PSII and PBSs was mapped making use of their fluorescence kinetics. Both radial and lateral heterogeneity were found in the distribution of the photosystems. State transitions were studied at the level of single cells. FLIM results show that PSII quenching occurs in all cells, irrespective of their state (I or II). In S. elongatus cells, this quenching is enhanced in State II. Furthermore, the decrease of PSII fluorescence in State II was homogeneous throughout the cells, despite the inhomogeneous PSI/PSII ratio. Finally, some disconnected PBSs were resolved in most State II cells. Taken together our data show that PSI is enriched in the inner thylakoid, while state transitions occur homogeneously throughout the cell.

scholarly journals The biochemical resolving power of fluorescence lifetime imaging

2021 ◽  
Author(s):  
Andrew L. Trinh ◽  
Alessandro Esposito
Keyword(s):  
Fluorescence Lifetime ◽  
Resolving Power ◽  
Fluorescence Lifetime Imaging ◽  
Time Resolved ◽  
Instrument Response Function ◽  
Lifetime Imaging ◽  
Use Of Time ◽  
Single Living Cells ◽  
Microscopy Techniques

AbstractA deeper understanding of spatial resolution in microscopy fostered a technological revolution that is now permitting us to investigate the structure of the cell with nanometer resolution. Although fluorescence microscopy techniques enable scientists to investigate both the structure and biochemistry of the cell, the biochemical resolving power of a microscope is a physical quantity that is not well-defined or studied. To overcome this limitation, we carried out a theoretical investigation of the biochemical resolving power in fluorescence lifetime imaging microscopy, one of the most effective tools to investigate biochemistry in single living cells. With the theoretical analysis of information theory and Monte Carlo simulations, we describe how the ‘biochemical resolving power’ in time-resolved sensing depends on instrument specifications. We unravel common misunderstandings on the role of the instrument response function and provide theoretical insights that have significant practical implications in the design and use of time-resolved instrumentation.

A Time-Resolved Four-Tap Lock-In Pixel CMOS Image Sensor for Real-Time Fluorescence Lifetime Imaging Microscopy

2018 ◽  
Vol 53 (8) ◽  
pp. 2319-2330 ◽  
Author(s):  
Min-Woong Seo ◽  
Yuya Shirakawa ◽  
Yoshimasa Kawata ◽  
Keiichiro Kagawa ◽  
Keita Yasutomi ◽  
...  
Keyword(s):  
Real Time ◽  
Fluorescence Lifetime ◽  
Cmos Image Sensor ◽  
Image Sensor ◽  
Fluorescence Lifetime Imaging Microscopy ◽  
Fluorescence Lifetime Imaging ◽  
Time Resolved ◽  
Lifetime Imaging ◽  
Lock In

Confocal fluorescence lifetime imaging of free calcium in single cells

1994 ◽  
Vol 4 (4) ◽  
pp. 291-294 ◽  
Author(s):  
Renata Sanders ◽  
Hans C. Gerritsen ◽  
Arie Draaijer ◽  
Piet M. Houpt ◽  
Yehudi K. Levine
Keyword(s):  
Fluorescence Lifetime ◽  
Single Cells ◽  
Free Calcium ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Imaging ◽  
Confocal Fluorescence

scholarly journals A biosensor of local kinesin activity reveals roles of PKC and EB1 in KIF17 activation

2013 ◽  
Vol 203 (3) ◽  
pp. 445-455 ◽  
Author(s):  
Cedric Espenel ◽  
Bipul R. Acharya ◽  
Geri Kreitzer
Keyword(s):  
Energy Transfer ◽  
Fluorescence Lifetime ◽  
Single Cells ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Epithelial Differentiation ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Data ◽  
Lifetime Imaging ◽  
Phasor Plot

We showed previously that the kinesin-2 motor KIF17 regulates microtubule (MT) dynamics and organization to promote epithelial differentiation. How KIF17 activity is regulated during this process remains unclear. Several kinesins, including KIF17, adopt compact and extended conformations that reflect autoinhibited and active states, respectively. We designed biosensors of KIF17 to monitor its activity directly in single cells using fluorescence lifetime imaging to detect Förster resonance energy transfer. Lifetime data are mapped on a phasor plot, allowing us to resolve populations of active and inactive motors in individual cells. Using this biosensor, we demonstrate that PKC contributes to the activation of KIF17 and that this is required for KIF17 to stabilize MTs in epithelia. Furthermore, we show that EB1 recruits KIF17 to dynamic MTs, enabling its accumulation at MT ends and thus promoting MT stabilization at discrete cellular domains.

scholarly journals TIME-RESOLVED IMAGING SPECTROSCOPY OF PLANT ADAPTATIONS TO CHANGES IN THE LIGHT ENVIRONMENT AND APPLICABILITY TO SCREENING MUTANTS

HortScience ◽  
1994 ◽  
Vol 29 (4) ◽  
pp. 249b-249
Author(s):  
Sylvain L. Dubé ◽  
John F. Allen
Keyword(s):  
Energy Balance ◽  
Physiological State ◽  
Imaging Spectroscopy ◽  
State Transitions ◽  
Time Resolved ◽  
Growth And Survival ◽  
Plant Adaptations ◽  
Physico Chemical ◽  
Time Resolved Imaging ◽  
Kinetics Of

Photosynthesis, a major determinant in growth and survival of plants, is very sensitive to the energy balance of the processes triggered by the physico-chemical environment. It is, therefore, an excellent indicator of the plants' physiological state. Fundamental events in photosynthesis can be studied non-invasively and non-destructively by examining there-emission of absorbed light energy as chlorophyll a fluorescence. In this study we present digitized consecutive images of fluorescence of intact leaves of Arabidopsis sp. The relative intensity and kinetics of fluorescence of several AOI (areas of interests) of each image have been analyzed and compared. We demonstrate the feasibility of this technique for studying the physiology of light adaptations (state-transitions) of several organisms simultaneously and its applicability in indentifying mutants. Implications of this technique to the horticulture industry will be discussed.

scholarly journals Fast fit-free analysis of fluorescence lifetime imaging via deep learning

2019 ◽  
Vol 116 (48) ◽  
pp. 24019-24030 ◽  
Author(s):  
Jason T. Smith ◽  
Ruoyang Yao ◽  
Nattawut Sinsuebphon ◽  
Alena Rudkouskaya ◽  
Nathan Un ◽  
...  
Keyword(s):  
Deep Learning ◽  
Fluorescence Lifetime ◽  
Near Infrared ◽  
Quantitative Information ◽  
Fluorescence Lifetime Imaging ◽  
Complex Data ◽  
Lifetime Imaging ◽  
Spatially Resolved ◽  
Lifetime Studies ◽  
Net Framework

Fluorescence lifetime imaging (FLI) provides unique quantitative information in biomedical and molecular biology studies but relies on complex data-fitting techniques to derive the quantities of interest. Herein, we propose a fit-free approach in FLI image formation that is based on deep learning (DL) to quantify fluorescence decays simultaneously over a whole image and at fast speeds. We report on a deep neural network (DNN) architecture, named fluorescence lifetime imaging network (FLI-Net) that is designed and trained for different classes of experiments, including visible FLI and near-infrared (NIR) FLI microscopy (FLIM) and NIR gated macroscopy FLI (MFLI). FLI-Net outputs quantitatively the spatially resolved lifetime-based parameters that are typically employed in the field. We validate the utility of the FLI-Net framework by performing quantitative microscopic and preclinical lifetime-based studies across the visible and NIR spectra, as well as across the 2 main data acquisition technologies. These results demonstrate that FLI-Net is well suited to accurately quantify complex fluorescence lifetimes in cells and, in real time, in intact animals without any parameter settings. Hence, FLI-Net paves the way to reproducible and quantitative lifetime studies at unprecedented speeds, for improved dissemination and impact of FLI in many important biomedical applications ranging from fundamental discoveries in molecular and cellular biology to clinical translation.

scholarly journals Synthesis, Photophysics, and Solvatochromic Studies of an Aggregated-Induced-Emission Luminogen Useful in Bioimaging

Sensors ◽  
2019 ◽  
Vol 19 (22) ◽  
pp. 4932 ◽  
Author(s):  
Laura Espinar-Barranco ◽  
Marta Meazza ◽  
Azahara Linares-Perez ◽  
Ramon Rios ◽  
Jose Manuel Paredes ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Fluorescence Emission ◽  
Fluorescence Lifetime Imaging ◽  
Selective Isolation ◽  
Time Resolved Fluorescence ◽  
Time Resolved ◽  
Lifetime Imaging ◽  
Intracellular Compartments ◽  
Photophysical Behavior ◽  
Heterogeneous Matrix

Biological samples are a complex and heterogeneous matrix where different macromolecules with different physicochemical parameters cohabit in reduced spaces. The introduction of fluorophores into these samples, such as in the interior of cells, can produce changes in the fluorescence emission properties of these dyes, caused by the specific physicochemical properties of cells. This effect can be especially intense with solvatofluorochromic dyes, where changes in the polarity environment surrounding the dye can drastically change the fluorescence emission. In this article, we studied the photophysical behavior of a new dye and confirmed the aggregation-induced emission (AIE) phenomenon with different approaches, such as by using different solvent proportions, increasing the viscosity, forming micelles, and adding bovine serum albumin (BSA), through analysis of the absorption and steady-state and time-resolved fluorescence. Our results show the preferences of the dye for nonpolar media, exhibiting AIE under specific conditions through immobilization. Additionally, this approach offers the possibility of easily determining the critical micelle concentration (CMC). Finally, we studied the rate of spontaneous incorporation of the dye into cells by fluorescence lifetime imaging and observed the intracellular pattern produced by the AIE. Interestingly, different intracellular compartments present strong differences in fluorescence intensity and fluorescence lifetime. We used this difference to isolate different intracellular regions to selectively study these regions. Interestingly, the fluorescence lifetime shows a strong difference in different intracellular compartments, facilitating selective isolation for a detailed study of specific organelles.

Sign in / Sign up

Export Citation Format

Share Document