Dynamic image correlation spectroscopy (ICS) and two-color image cross-correlation spectroscopy (ICCS): concepts and application

2000 ◽  
Author(s):  
Paul W. Wiseman ◽  
Jeffrey A. Squier ◽  
Kent R. Wilson
Keyword(s):  
Cross Correlation ◽  
Color Image ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Dynamic Image ◽  
Image Correlation Spectroscopy

Two-photon image correlation spectroscopy and image cross-correlation spectroscopy

2000 ◽  
Vol 200 (1) ◽  
pp. 14-25 ◽  
Author(s):  
P. W. Wiseman ◽  
J. A. Squier ◽  
M. H. Ellisman ◽  
K. R. Wilson
Keyword(s):  
Cross Correlation ◽  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Two Photon ◽  
Image Correlation Spectroscopy

Exploring oligomeric state of the serotonin1A receptor utilizing photobleaching image correlation spectroscopy: implications for receptor function

2018 ◽  
Vol 207 ◽  
pp. 409-421 ◽  
Author(s):  
Hirak Chakraborty ◽  
Md. Jafurulla ◽  
Andrew H. A. Clayton ◽  
Amitabha Chattopadhyay
Keyword(s):  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Membrane Cholesterol ◽  
Receptor Function ◽  
Oligomeric State ◽  
Serotonin1a Receptor ◽  
Image Correlation Spectroscopy

Photobleaching image correlation spectroscopy (pbICS) reveals that membrane cholesterol modulates the oligomeric state of the serotonin1A receptor.

scholarly journals Arbitrary-Region Image Correlation Spectroscopy

2016 ◽  
Vol 110 (3) ◽  
pp. 176a
Author(s):  
Jelle Hendrix ◽  
Tomas Dekens ◽  
Don C. Lamb
Keyword(s):  
Correlation Spectroscopy ◽  
Image Correlation ◽  
Arbitrary Region ◽  
Image Correlation Spectroscopy

scholarly journals Visualization of class A GPCR oligomerization by image-based fluorescence fluctuation spectroscopy

2017 ◽  
Author(s):  
Ali Isbilir ◽  
Jan Möller ◽  
Andreas Bock ◽  
Ulrike Zabel ◽  
Paolo Annibale ◽  
...  
Keyword(s):  
Single Molecule ◽  
Metabotropic Glutamate Receptors ◽  
Fluorescent Protein ◽  
Methodological Approach ◽  
Correlation Spectroscopy ◽  
Membrane Receptors ◽  
Image Correlation ◽  
Intact Cells ◽  
Class A ◽  
Image Correlation Spectroscopy

AbstractG protein-coupled receptors (GPCRs) represent the largest class of cell surface receptors conveying extracellular information into intracellular signals. Many GPCRs have been shown to be able to oligomerize and it is firmly established that Class C GPCRs (e.g. metabotropic glutamate receptors) function as obligate dimers. However, the oligomerization capability of the larger Class A GPCRs (e.g. comprising the β-adrenergic receptors (β-ARs)) is still, despite decades of research, highly debated.Here we assess the oligomerization behavior of three prototypical Class A GPCRs, the β1-ARs, β2-ARs, and muscarinic M2Rs in single, intact cells. We combine two image correlation spectroscopy methods based on molecular brightness, i.e. the analysis of fluorescence fluctuations over space and over time, and thereby provide an assay able to robustly and precisely quantify the degree of oligomerization of GPCRs. In addition, we provide a comparison between two labelling strategies, namely C-terminally-attached fluorescent proteins and N-terminally-attached SNAP-tags, in order to rule out effects arising from potential fluorescent protein-driven oligomerization. The degree of GPCR oligomerization is expressed with respect to a set of previously reported as well as newly established monomeric or dimeric control constructs. Our data reveal that all three prototypical GPRCs studied display, under unstimulated conditions, a prevalently monomeric fingerprint. Only the β2-AR shows a slight degree of oligomerization.From a methodological point of view, our study suggests three key aspects. First, the combination of two image correlation spectroscopy methods allows addressing cells transiently expressing high concentrations of membrane receptors, far from the single molecule regime, at a density where the kinetic equilibrium should favor dimers and higher-order oligomers. Second, our methodological approach, allows to selectively target cell membrane regions devoid of artificial oligomerization hot-spots (such as vesicles). Third, our data suggest that the β1-AR appears to be a superior monomeric control than the widely used membrane protein CD86.Taken together, we suggest that our combined image correlation spectroscopy method is a powerful approach to assess the oligomerization behavior of GPCRs in intact cells at high expression levels.

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