University of Würzburg, Germany

2 ARTICLES PUBLISHED IN JoVE

Bioengineering

High-resolution Spatiotemporal Analysis of Receptor Dynamics by Single-molecule Fluorescence Microscopy

Titiwat Sungkaworn¹, Finn Rieken¹, Martin J. Lohse¹, Davide Calebiro¹

¹Institute of Pharmacology and Toxicology and Bio-Imaging Center/Rudolf Virchow Center, DFG-Research Center for Experimental Biomedicine, University of Würzburg, Germany

This protocol describes how to use total internal reflection fluorescence microscopy to visualize and track single receptors on the surface of living cells and thereby analyze receptor lateral mobility, size of receptor complexes as well as to visualize transient receptor-receptor interactions. This protocol can be extended to other membrane proteins.

Biochemistry

Dual-Color Fluorescence Cross-Correlation Spectroscopy to Study Protein-Protein Interaction and Protein Dynamics in Live Cells

Katherina Hemmen^*1, Susobhan Choudhury^*1, Mike Friedrich¹, Johannes Balkenhol¹, Felix Knote¹, Martin J. Lohse², Katrin G. Heinze¹

¹Rudolf Virchow Center for Integrative and Translation Bioimaging, Julius-Maximilian University Wuerzburg, ²Max Delbrück Center for Molecular Medicine, Berlin

We present an experimental protocol and data analysis workflow to perform live cell dual-color fluorescence cross correlation spectroscopy (FCCS) combined with Förster Resonance Energy transfer (FRET) to study membrane receptor dynamics in live cells using modern fluorescence labeling techniques.