An experimental setup for femtosecond laser modification and confocal spectroscopy of bioloigcal tissue

An experimental setup was designed and built to spectroscopically investigate biological tissues. Described is a confocal microscope setup built to perform both linear and nonlinear laser spectroscopy as well as femtosecond laser modifications of biological tissues in a combined setup. Of interest was corneal tissue modified by femtosecond laser pulses at intensities below the laser induced optical breakdown threshold. Tissue modifications, which appear as streaks, indicate that changes are taking place in the molecular structure of the tissue despite the lack of full optical breakdown. These modifications are of interest because of the expanding use of femtosecond lasers in refractive surgery. In this work, the design of an experimental setup capable of performing several types of linear and nonlinear spectroscopic methods applicable to studying these modifications is described. The confocal spectroscopy setup consists of several excitation lasers, a spectrometer and detectors for acquiring spectra, and a laser-scanning stage for spectral imaging. In addition, the setup integrates the ability to modify corneal tissue by femtosecond laser exposure so that modifications can be performed and monitored in the same experimental apparatus. Experiments were performed to attempt corneal tissue modifications by femtosecond laser. Modifications were successful using an amplified Ti:Sapphire laser with ∼100 nJ pulses. To test the spectroscopic capabilities of the setup, Raman and fluorescence spectroscopy experiments were done on ocular tissues from freshly enucleated lamb eyes. Spectra of lens and corneal tissue were acquired using 660 nm and 473 nm diode lasers. The setup and data acquired by these preliminary experiments will establish a basis for a future in depth investigation of these corneal modifications.