scholarly journals Monitoring Dynamic Binding of Chromatin Proteins In Vivo by Fluorescence Correlation Spectroscopy and Temporal Image Correlation Spectroscopy

2011 ◽  
pp. 177-200 ◽  
Author(s):  
Davide Mazza ◽  
Timothy J. Stasevich ◽  
Tatiana S. Karpova ◽  
James G. McNally
Keyword(s):  
Fluorescence Correlation Spectroscopy ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Dynamic Binding ◽  
Fluorescence Correlation ◽  
Chromatin Proteins ◽  
Image Correlation Spectroscopy

Investigating membrane protein dynamics in living cellsThis paper is one of a selection of papers published in this Special Issue, entitled CSBMCB — Membrane Proteins in Health and Disease.

2006 ◽  
Vol 84 (6) ◽  
pp. 825-831 ◽  
Author(s):  
Ian R. Bates ◽  
Paul W. Wiseman ◽  
John W. Hanrahan
Keyword(s):  
Membrane Proteins ◽  
Fluorescence Correlation Spectroscopy ◽  
Fluorescence Recovery After Photobleaching ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Fluorescence Correlation ◽  
Transport Dynamics ◽  
Health And Disease ◽  
Image Correlation Spectroscopy ◽  
Selection Of

Live cell imaging is a powerful tool for understanding the function and regulation of membrane proteins. In this review, we briefly discuss 4 fluorescence-microscopy-based techniques for studying the transport dynamics of membrane proteins: fluorescence-correlation spectroscopy, image-correlation spectroscopy, fluorescence recovery after photobleaching, and single-particle and (or) molecule tracking. The advantages and limitations of each approach are illustrated using recent studies of an ion channel and cell adhesion molecules.

Active protein transport through plastid tubules: velocity quantified by fluorescence correlation spectroscopy

2000 ◽  
Vol 113 (22) ◽  
pp. 3921-3930 ◽  
Author(s):  
R.H. Kohler ◽  
P. Schwille ◽  
W.W. Webb ◽  
M.R. Hanson
Keyword(s):  
Active Transport ◽  
Fluorescence Correlation Spectroscopy ◽  
Protein Transport ◽  
Fluorescent Protein ◽  
Correlation Spectroscopy ◽  
Long Distance ◽  
Fluorescence Correlation ◽  
Photon Excitation ◽  
Green Fluorescent

Dynamic tubular projections emanate from plastids in certain cells of vascular plants and are especially prevalent in non-photosynthetic cells. Tubules sometimes connect two or more different plastids and can extend over long distances within a cell, observations that suggest that the tubules may function in distribution of molecules within, to and from plastids. In a new application of two-photon excitation (2PE) fluorescence correlation spectroscopy (FCS), we separated diffusion of fluorescent molecules from active transport in vivo. We quantified the velocities of diffusion versus active transport of green fluorescent protein (GFP) within plastid tubules and in the cytosol in vivo. GFP moves by 3-dimensional (3-D) diffusion both in the cytosol and plastid tubules, but diffusion in tubules is about 50 times and 100 times slower than in the cytosol and an aqueous solution, respectively. Unexpectedly larger GFP units within plastid tubules exhibited active transport with a velocity of about 0.12 microm/second. Active transport might play an important role in the long-distance distribution of large numbers of molecules within the highly viscous stroma of plastid tubules.

scholarly journals Fluctuations as a source of information in fluorescence microscopy

2008 ◽  
Vol 6 (suppl_1) ◽  
Author(s):  
Zdeněk Petrášek ◽  
Petra Schwille
Keyword(s):  
Single Molecule ◽  
Single Point ◽  
Quantitative Information ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Photon Detection ◽  
Fluorescence Correlation ◽  
Additional Information ◽  
Image Correlation Spectroscopy ◽  
Spatio Temporal

Fluctuations in fluorescence spectroscopy and microscopy have traditionally been regarded as noise—they lower the resolution and contrast and do not permit high acquisition rates. However, fluctuations can also be used to gain additional information about a system. This fact has been exploited in single-point microscopic techniques, such as fluorescence correlation spectroscopy and analysis of single molecule trajectories, and also in the imaging field, e.g. in spatio-temporal image correlation spectroscopy. Here, we discuss how fluctuations are used to obtain more quantitative information from the data than that given by average values, while minimizing the effects of noise due to stochastic photon detection.

scholarly journals In vivo fluorescence correlation spectroscopy analyses of FMBP ‐1, a silkworm transcription factor

FEBS Open Bio ◽  
2016 ◽  
Vol 6 (2) ◽  
pp. 106-125 ◽  
Author(s):  
Motosuke Tsutsumi ◽  
Hideki Muto ◽  
Shohei Myoba ◽  
Mai Kimoto ◽  
Akira Kitamura ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Fluorescence Correlation Spectroscopy ◽  
Correlation Spectroscopy ◽  
Fluorescence Correlation ◽  
In Vivo Fluorescence

scholarly journals Protein-Protein Interactions In Vivo Studied by Single Plane Illumination Fluorescence Correlation Spectroscopy (SPIM-FCS)

2013 ◽  
Vol 104 (2) ◽  
pp. 575a ◽  
Author(s):  
Agata Pernus ◽  
Jan Krieger ◽  
Jan Buchholz ◽  
Anand Pratap Singh ◽  
Edoardo Charbon ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Fluorescence Correlation Spectroscopy ◽  
Correlation Spectroscopy ◽  
Protein Protein Interactions ◽  
Single Plane ◽  
Fluorescence Correlation

scholarly journals Antenna Protein Clustering In Vitro Unveiled by Fluorescence Correlation Spectroscopy

2021 ◽  
Vol 22 (6) ◽  
pp. 2969
Author(s):  
Aurélie Crepin ◽  
Edel Cunill-Semanat ◽  
Eliška Kuthanová Trsková ◽  
Erica Belgio ◽  
Radek Kaňa
Keyword(s):  
Fluorescence Correlation Spectroscopy ◽  
Higher Plants ◽  
Fluorescence Emission ◽  
Correlation Spectroscopy ◽  
Photochemical Quenching ◽  
High Ph ◽  
Fluorescence Correlation ◽  
Non Photochemical Quenching

Antenna protein aggregation is one of the principal mechanisms considered effective in protecting phototrophs against high light damage. Commonly, it is induced, in vitro, by decreasing detergent concentration and pH of a solution of purified antennas; the resulting reduction in fluorescence emission is considered to be representative of non-photochemical quenching in vivo. However, little is known about the actual size and organization of antenna particles formed by this means, and hence the physiological relevance of this experimental approach is questionable. Here, a quasi-single molecule method, fluorescence correlation spectroscopy (FCS), was applied during in vitro quenching of LHCII trimers from higher plants for a parallel estimation of particle size, fluorescence, and antenna cluster homogeneity in a single measurement. FCS revealed that, below detergent critical micelle concentration, low pH promoted the formation of large protein oligomers of sizes up to micrometers, and therefore is apparently incompatible with thylakoid membranes. In contrast, LHCII clusters formed at high pH were smaller and homogenous, and yet still capable of efficient quenching. The results altogether set the physiological validity limits of in vitro quenching experiments. Our data also support the idea that the small, moderately quenching LHCII oligomers found at high pH could be relevant with respect to non-photochemical quenching in vivo.

Sign in / Sign up

Export Citation Format

Share Document