Fiber Based Non-linear Excitation Fluorescence Microscopy

Abstract

Non-linear excitation fluorescence microscopy is a high resolution, non-invasive biological imaging technique capable of imaging both in vitro and in vivo at the depths of several hundred microns. The goal of the thesis work is to contribute towards a fiber based non-linear excitation fluorescence microscopy system that we plan to build for our lab, which would later on be integrated with the OCT imaging setup and the DIC microscope. This thesis work involves filling in the key components in the microscopy system. These key components are; femtosecond pulse diagnosis, study of dispersion in optical media, dispersion free femtosecond pulse delivery and design and construction of an experimental multi-photon imaging setup which would be used for imaging biological samples. An interferometric autocorrelator was designed, built and interfaced with the computer for diagnosis of femtosecond laser pulses. In order to quantify the dispersion in optical fibers which are common choice for flexible delivery of light, a mathematical simulation was created. The results from the simulation indicated that optical fibers could not be used as the means to deliver ultrashort laser pulses for non-linear excitation fluorescence microscopy application. For the purpose of flexible delivery of ultrashort pulses as well as preserving their ultrashort temporal profile, hollow core photonic bandgap fiber (PCF) was chosen and characterized. In the last part of the thesis work, an experimental setup of a two-photon excitation fluorescence microscope was designed and built with a flexible delivery setup and laser beam diagnosis setup.