Practical two-photon-absorption cross sections and spectra of eosin and hematoxylin

We experimentally investigate the two-photon absorption cross sections and spectra of eosin and hematoxylin for applications in nonlinear microscopy. The experiments are carried out on pure samples of the two dyes in DI-water solvent with different concentrations, in the typical range employed in standard staining procedures. Nonlinear fluorescence is excited by a line-shaped beam emitted by a Ti:Sapphire mode-locked laser in the wavelength range from 740 to 880 nm and is detected through a microscope setup. The two-photon absorption spectral response is systematically analyzed and discussed. Finally, the staining is applied on biological tissue samples, which are imaged by two-photon microscopy. Our results show that the employed dyes are fully suitable for applications in nonlinear imaging.