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Investigatation of the photo-induced transient states of Merocyanine 540 (MC540) by Fluorescence correlation spectroscopy (FCS) (CAT#: STEM-MB-1168-WXH)

Introduction

Merocyanines are a class of polymethine dyes which are clearly defined by set structural properties. Merycyanines belong to the group of dyes referred to as functional dyes, where their applications are not only determined by their colour, but also their valuable chemical properties.These dyes are usually intensely colored and have large extinction coefficients.
Merocyanine 540 was the first fluorescent dye used for measuring membrane potential.




Principle

Fluorescence correlation spectroscopy (FCS) is a statistical analysis, via time correlation, of stationary fluctuations of the fluorescence intensity. Its theoretical underpinning originated from L. Onsager's regression hypothesis. The analysis provides kinetic parameters of the physical processes underlying the fluctuations. One of the interesting applications of this is an analysis of the concentration fluctuations of fluorescent particles (molecules) in solution. In this application, the fluorescence emitted from a very tiny space in solution containing a small number of fluorescent particles (molecules) is observed. The fluorescence intensity is fluctuating due to Brownian motion of the particles. In other words, the number of the particles in the sub-space defined by the optical system is randomly changing around the average number. The analysis gives the average number of fluorescent particles and average diffusion time, when the particle is passing through the space. Eventually, both the concentration and size of the particle (molecule) are determined. Both parameters are important in biochemical research, biophysics, and chemistry.

Applications

• Measurement of the diffusion coefficient of biomolecules
• Detection of translational diffusions
• Measurement of the biomolecular concentration in vitro or in vivo
• Quantification of the viscosity of a solution
• Monitoring the binding or unbinding of two kinds of biomolecules
• Probing the diffusion paths of different directions and mapping the intercellular obstacles

Procedure

1. Sample Preparation
2. Fluorescence correlation spectroscopy (FCS) testing
3. Data analysis

Materials

Fluorescence Correlation Spectrometer
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