Fluorescence and light diffusion

There are two types of light diffusion Rayleigh and Raman. Energies of diffused and excited photons are equal. Thus, Rayleigh diffusion and excitation occurs at the same wavelengths. 

There are two types of light difhision Rayleigh and Raman. Rayleigh diffusion occurs at wavelength equal to that of the excitation one. Thus, energy of the diffused photons is equal to the energy of the excitation photons. 

Usually one should not start the emission spectrum before or at the excitation wavelength in order to avoid recording Rayleigh diffusion. 

Raman diffusion is observed at a wavelength that is higher than the excitation wavelength. The diffused photons have quantum energy inferior to that of the excitation photons. Therefore, the intensity of the Raman peak is lower than that of the Rayleigh peak. Raman can perturb the emission spectrum and thus one should subtract the Raman from the emission spectrum. 

In general, more the fluorescence intensity is important, less intense will be the Raman peak. In aqueous medium, the 0-H bonds are responsible for the Raman spectrum. The position of the Raman peak is dependent on the excitation wavelength. It is possible to calculate the position of the Raman peak in aqueous medium with the following equation  

Fluorophores are spread in nature and thus ih can be used as natural indicators to study the structure, the dynamics and the metabolism of living cells. 

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