TY - CHAP
T1 - Chapter 2 Molecular Sensors Based on Fluorescence Resonance Energy Transfer to Visualize Cellular Dynamics
AU - Ananthanarayanan, Bharath
AU - Ni, Qiang
AU - Zhang, Jin
N1 - Funding Information:
Work of the authors was supported by the National Institutes of Health (DK073368 and CA122673 to J. Z.), a Scientist Development Award from the American Heart Association (to J. Z.), a Young Clinical Scientist Award from Flight Attendant Medical Institute (to J. Z.), 3M (to J. Z.), and the W.M. Keck Center (to J. Z. and B. A.).
PY - 2008
Y1 - 2008
N2 - Visualizing a variety of signaling events in the native cellular environment is now possible with the advent of genetically encodable fluorescent labels like green fluorescent proteins made de novo by living cells themselves. The focus of this method chapter is on genetically encodable molecular sensors based on fluorescence resonance energy transfer (FRET) for visualization of cellular dynamics. This chapter discusses the process of developing a molecular sensor, from choosing donor-acceptor pairs to designing the protein modules that actually sense the signaling events. A few examples of biosensors are discussed to showcase the designs of such FRET-based sensors for live-cell imaging of signaling events. Subsequently, Section III covers the experimental procedure of DNA work, microscope instrumentation, data collection through imaging acquisition, data comprehension, and evaluation. Furthermore, a case study of the PI3K/Akt signaling pathway using a series of FRET sensors highlights the tremendous potential of the method in exploring relevant biological systems.
AB - Visualizing a variety of signaling events in the native cellular environment is now possible with the advent of genetically encodable fluorescent labels like green fluorescent proteins made de novo by living cells themselves. The focus of this method chapter is on genetically encodable molecular sensors based on fluorescence resonance energy transfer (FRET) for visualization of cellular dynamics. This chapter discusses the process of developing a molecular sensor, from choosing donor-acceptor pairs to designing the protein modules that actually sense the signaling events. A few examples of biosensors are discussed to showcase the designs of such FRET-based sensors for live-cell imaging of signaling events. Subsequently, Section III covers the experimental procedure of DNA work, microscope instrumentation, data collection through imaging acquisition, data comprehension, and evaluation. Furthermore, a case study of the PI3K/Akt signaling pathway using a series of FRET sensors highlights the tremendous potential of the method in exploring relevant biological systems.
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U2 - 10.1016/S0091-679X(08)00602-X
DO - 10.1016/S0091-679X(08)00602-X
M3 - Chapter
C2 - 19118671
AN - SCOPUS:57949100104
SN - 9780123725219
T3 - Methods in Cell Biology
SP - 37
EP - 57
BT - Biophysical Tools for Biologists, Volume Two
A2 - Correia, John
A2 - Detrich III, William
ER -