This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Quantifying membrane protein oligomerization using two-dimensional fluorescence intensity fluctuation spectrometry
Valerică Raicu
Physics Department, University of Wisconsin-Milwaukee
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Current methods for determining membrane protein association in cells are either very time consuming, require complicated procedures, or lack the sensitivity needed to assess the effect of concentration or ligand binding on the observed oligomerization. To overcome these limitations, we provide a detailed protocol for quantifying the relative abundance and stability of oligomers of differing sizes using two-dimensional fluorescence intensity fluctuation (2D-FIF) spectrometry. The approach can be implemented using a standard laser scanning fluorescence microscope along with custom written software for image analysis. This method may be applied to evaluate the extent of oligomerization as a function of receptor concentration and is particularly suited to assess the effects of agonists and antagonists on the oligomeric size of membrane receptors of interest.
Keywords
Fluorescence microscopy, Fluorescence fluctuation spectroscopy, G protein-coupled receptor, protein-protein interaction, oligomerizatioin, quaternary structure
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 11 Jul, 2019
No community comments so far
Metrics
Comments: 0
HTML Views: ...
Subject Areas
Biological techniques
Learn more about our company.
This is a preprint. It has not completed peer review.
Quantifying membrane protein oligomerization using two-dimensional fluorescence intensity fluctuation spectrometry
Valerică Raicu
Physics Department, University of Wisconsin-Milwaukee
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 11 Jul, 2019
No community comments so far
Metrics
Comments: 0
HTML Views: ...
Subject Areas
Biological techniques
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Current methods for determining membrane protein association in cells are either very time consuming, require complicated procedures, or lack the sensitivity needed to assess the effect of concentration or ligand binding on the observed oligomerization. To overcome these limitations, we provide a detailed protocol for quantifying the relative abundance and stability of oligomers of differing sizes using two-dimensional fluorescence intensity fluctuation (2D-FIF) spectrometry. The approach can be implemented using a standard laser scanning fluorescence microscope along with custom written software for image analysis. This method may be applied to evaluate the extent of oligomerization as a function of receptor concentration and is particularly suited to assess the effects of agonists and antagonists on the oligomeric size of membrane receptors of interest.