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Related Concept Videos

Atomic Fluorescence Spectroscopy01:29

Atomic Fluorescence Spectroscopy

Atomic fluorescence spectroscopy (AFS) is an analytical technique that involves the electronic transitions of atoms in a flame, furnace, or plasma being excited by electromagnetic (EM) radiation. When these atoms absorb energy, they become excited and subsequently release energy as they return to their original state. This emitted light, or "fluorescence," is observed at a right angle to the incident beam. Both absorption and emission processes transpire at distinct wavelengths, which are...
Fluorescence and Phosphorescence: Instrumentation01:25

Fluorescence and Phosphorescence: Instrumentation

Fluorometers and spectrofluorometers are two types of instruments used for measuring molecular fluorescence. These instruments differ in how they select excitation and emission wavelengths and the type of light sources they utilize. Fluorometers use absorption interference filters to choose excitation and emission wavelengths. The excitation source in a fluorometer is typically a low-pressure mercury vapor lamp that emits intense lines distributed throughout the ultraviolet and visible regions.
Confocal Fluorescence Microscopy01:16

Confocal Fluorescence Microscopy

Confocal microscopy is an advanced microscopic technique. The prime advantage of the confocal microscope over other microscopy techniques is its ability to block the out-of-focus light from the illuminated samples using pinholes. It is widely used with fluorescence optics to obtain high-resolution, sharp contrast images. Unlike optical microscopes, confocal microscopes use a focused beam of light laser to scan the entire sample surface at different z-planes. These microscopes are, therefore,...
Total Internal Reflection Fluorescence Microscopy01:05

Total Internal Reflection Fluorescence Microscopy

Total internal reflection fluorescence microscopy or TIRF is an advanced microscopic technique used to visualize fluorophores in samples close to a solid surface with a higher refractive index, such as a glass coverslip. TIRF only allows fluorophores in proximity to the solid surface to be excited. When light from a medium with a lower refractive index (such as air) hits the glass coverslip at a critical angle, the light undergoes total internal reflection stead of passing through the glass.
Super-resolution Fluorescence Microscopy01:37

Super-resolution Fluorescence Microscopy

Super-resolution fluorescence microscopy (SRFM) provides a better resolution than conventional fluorescence microscopy by reducing the point spread function (PSF). PSF is the light intensity distribution from a point that causes it to appear blurred. Due to PSF, each fluorescing point appears bigger than its actual size, and it is the PSF interference of nearby fluorophores that causes the blurred image. Various approaches to achieving higher resolution through SRFM have recently been developed.
Variables Affecting Phosphorescence and Fluorescence01:26

Variables Affecting Phosphorescence and Fluorescence

Fluorescence and phosphorescence are essential phenomena in fields like analytical chemistry, biological imaging, and materials science, where they detect molecular properties and visualize cellular structures. Understanding the variables that influence these luminescent behaviors is crucial for maximizing accuracy and efficiency in their applications. These variables can broadly be grouped into chemical structure, solvent properties, and external conditions, each playing a distinct role in...

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Updated: May 29, 2026

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

Published on: December 11, 2021

Fluorescence correlation spectroscopy.

Patrick Ferrand1, Jérôme Wenger, Hervé Rigneault

  • 1Mosaic Group, Institut Fresnel, CNRS, Aix-Marseille Université, Ecole Central Marseille, Marseille, France. patrick.ferrand@fresnel.fr

Methods in Molecular Biology (Clifton, N.J.)
|September 13, 2011
PubMed
Summary
This summary is machine-generated.

Fluorescence correlation spectroscopy (FCS) analyzes molecular dynamics by correlating intensity fluctuations in a confocal volume. This tutorial provides a practical guide to FCS principles, setup, and data analysis for researchers.

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Determination of Lipid Raft Partitioning of Fluorescently-tagged Probes in Living Cells by Fluorescence Correlation Spectroscopy (FCS)
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Area of Science:

  • Biophysics
  • Chemical Physics
  • Microscopy

Background:

  • Fluorescence correlation spectroscopy (FCS) is a powerful microscopy technique.
  • It analyzes time-fluctuating intensity from molecules diffusing through a confocal volume.
  • FCS quantifies molecular properties like diffusion, photophysics, and interactions.

Purpose of the Study:

  • To provide a practical, hands-on introduction to Fluorescence Correlation Spectroscopy (FCS).
  • To explain the core principles, theoretical assumptions, and analytical expressions of FCS.
  • To guide users in building, calibrating, measuring with, and analyzing data from FCS systems.

Main Methods:

  • Detailed explanation of the autocorrelation function for intensity fluctuations.
  • Discussion of critical parameters for constructing an FCS setup.
  • Step-by-step operational guide covering calibration, measurement, and data processing.

Main Results:

  • Emphasis on theoretical underpinnings and key parameters for FCS implementation.
  • Practical examples illustrating common challenges in FCS measurements.
  • Proposed solutions to overcome experimental difficulties in FCS.

Conclusions:

  • FCS is a versatile tool for studying molecular dynamics and properties.
  • Understanding theoretical principles and practical considerations is crucial for successful FCS experiments.
  • This tutorial aims to demystify FCS for new users, enabling effective application.