Raster Image Correlation Spectroscopy in Live cells Expressing Endothelin ETA Receptor

Order of Publishing in Issue: 
Volume :7
Issue :1
January, 2013
Page No: 
Damaris De La Torre [a,e], Elizabeth A. Gordon [b], Michelle A. Digman [b,c, Milka Stakicc, Hanns Häberlein [d], Enrico Gratton [bc] and Catherina Caballero-George [a]
[a] Unit of Molecular Pharmacology and Pharmacognosy, Institute for Scientific Research and High Technology Services, Bld. 219, City of Knowledge, Clayton, Panama Republic
[b] Department of Developmental and Cell Biology, University of California, Irvine, California, USA
[c] Laboratory for Fluorescence Dynamics, Department of Biomedical Engineering, University of California, Irvine, California, USA
[d] Institut für Biochemie und Molekularbiologie, Nussallee 11, 53115 Bonn, Germany
[e] Department of Biotechnology, Acharya Nagarjuna University, Guntur, India

Abstract : Fluorescence spectroscopy is the most common non-radioactive technique used to study GPCR interactions with their ligands. Raster image correlation spectroscopy (RICS) exploits spatio-temporal correlation functions rather than the simple temporal correlations of conventional fluorescence correlation spectroscopy. In this paper we describe the use of RICS and the number and brightness method to determine the diffusion of a construct of endothelin ETA receptor with EGFP and the aggregation state in the cytoplasm. Our construct seems to locate mainly in the cytoplasm where it undergoes diffusion and it appears to be monomeric. Although our construct could not fully represent the native protein, we believe that the methodology we describe in this paper could be used by anyone in this field.

Key words: Endothelin ETA receptor, GPCR, live cell, imaging, RICS.

Endothelin ETA receptor, GPCR, live cell, imaging, RICS.
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