Abstract
Non linear optical scanning microscopy has became a useful tool for tissue imaging. Biological tissues are highly scattering media and this lead to an exponentially attenuation of the excitation intensity as the light moves into the sample. While performing imaging of biological scattering tissues in non linear excitation regime, the localization of the maximum 2PE intensity was found to shift closer to the surface [1] and the 2PE imaging depth limit appears strongly limited by near surface fluorescence [2]. In this work we computed the illumination and the photobleaching distribution [3] for different scattering coefficients in order to characterize the effects induced by scattering. An experimental test has been carried out by imaging, with medium numerical aperture objective (N.A.=0.8), thick scattering fluorescent immobile sample (polyelectrolyte gel). Results confirm that in this conditions no photobleaching effects due to scattering occur close to the surface.[1] J. P. Ying et al, Appl. Opt. 38, (1999).[2] P. Theer J. Opt. Soc. Am. A. 23, (2006).[3] D.Mazza et al, Appl. Opt. 46 (2007).
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