NDLI: Confocal FLIM of Genetically Encoded FRET Sensors for Quantitative $Ca^{2+}$ Imaging

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Confocal FLIM of Genetically Encoded FRET Sensors for Quantitative $Ca^{2+}$ Imaging

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Sauer, Benjamin Tian, Qinghai Lipp, Peter Kaestner, Lars
Description	Country affiliation: Germany Author Affiliation: Sauer B ( Institute for Molecular Cell Biology and Research Center for Molecular Imaging and Screening, School of Medicine, Saarland University, 66421 Homburg/Saar, Germany.); Tian Q ( Institute for Molecular Cell Biology and Research Center for Molecular Imaging and Screening, School of Medicine, Saarland University, 66421 Homburg/Saar, Germany.); Lipp P ( Institute for Molecular Cell Biology and Research Center for Molecular Imaging and Screening, School of Medicine, Saarland University, 66421 Homburg/Saar, Germany.); Kaestner L ( Institute for Molecular Cell Biology and Research Center for Molecular Imaging and Screening, School of Medicine, Saarland University, 66421 Homburg/Saar, Germany.)
Abstract	Fluorescence lifetime imaging (FLIM) is a powerful imaging mode that can be combined with confocal imaging. Changes in the fluorescence decay time of a donor in an intramolecular Förster resonance energy transfer (FRET)-based biosensor provide intrinsic quantitative data. Here, we describe a protocol using both the $Ca^{2+}$ sensor TN-XL, which uses troponin C, as the $Ca^{2+}-sensing$ unit, and the FLIM technology based on time-correlated single-photon counting.
File Format	HTM / HTML
ISSN	19403402
Issue Number	12
Volume Number	2014
e-ISSN	15596095
Journal	Cold Spring Harbor Protocols
Language	English
Publisher	Cold Spring Harbor Laboratory Press
Publisher Date	2014-12-01
Publisher Place	United States
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Discipline Clinical Laboratory Techniques Biosensing Techniques Calcium Metabolism Fluorescence Resonance Energy Transfer Methods Genes, Reporter Microscopy, Confocal Microscopy, Fluorescence Hek293 Cells Humans Photons Transfection Journal Article Research Support, Non-u.s. Gov't
Content Type	Text
Resource Type	Article
Subject	Biochemistry, Genetics and Molecular Biology

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1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
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Protocol 1: Verifying the function and localization of genetically-encoded $Ca^{2+}$ sensors and converting FRET ratios to $Ca^{2+}$ concentrations

Advanced Fluorescence Microscopy Techniques—FRAP, FLIP, FLAP, FRET and FLIM

FLIM-FRET analysis using $Ca^{2+}$ sensors in HeLa cells

Measuring the in situs $K_{d}$ of a genetically-encoded $Ca^{2+}$ sensor

Imaging neuronal activity with genetically encoded calcium indicators.

Non fitting based FRET–FLIM analysis approaches applied to quantify protein–protein interactions in live cells

Signaling Microdomains in the Spotlight: Visualizing Compartmentalized Signaling Using Genetically Encoded Fluorescent Biosensors.

Genetically encoded sensors of protein hydrodynamics and molecular proximity

Properties of genetically-encoded indicators used for cytosolic and organellar $Ca^{2+}$ measurements

Confocal FLIM of Genetically Encoded FRET Sensors for Quantitative $Ca^{2+}$ Imaging

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Protocol 1: Verifying the function and localization of genetically-encoded $Ca^{2+}$ sensors and converting FRET ratios to $Ca^{2+}$ concentrations

Advanced Fluorescence Microscopy Techniques—FRAP, FLIP, FLAP, FRET and FLIM

FLIM-FRET analysis using $Ca^{2+}$ sensors in HeLa cells

Measuring the in situs $K_{d}$ of a genetically-encoded $Ca^{2+}$ sensor

Imaging neuronal activity with genetically encoded calcium indicators.

Non fitting based FRET–FLIM analysis approaches applied to quantify protein–protein interactions in live cells

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Confocal FLIM of Genetically Encoded FRET Sensors for Quantitative $Ca^{2+}$ Imaging