Improved fluorescence-enhanced optical imaging and tomography by enhanced excitation light rejection

Abstract

Fluorescence enhanced optical imaging and tomography studies involve the detection of weak fluorescent signals emanating from nano- to picomolar concentrations of exogenous or endogenously produced fluorophore concurrent with the rejection of an overwhelmingly large component of backscattered excitation light. The elimination of the back-reflected excitation light of the collected signal remains a major and often unrecognized challenge for further reducing the noise floor and increasing sensitivity of small animal fluorescence imaging. In this dissertation, we adapted collimating and gradient index (GRIN) lenses in an existing frequency-domain system to improve excitation light rejection and enhance planar and tomographic imaging. To achieve this goal, we developed planar and tomographic imaging systems based upon ray tracing calculations for improved rejection of excitation light. The “out-of-band (S (λx))” to “in-band (S (λm) - S (λx))” signal ratio assessing excitation leakage was acquired with and without collimating optics. The addition of collimating optics resulted in a 51 to 75% reduction in the transmission ratio of (S (λx))/ (S (λm) - S (λx)) for the phantom studies and an increase of target to background ratio (TBR) from 11% to 31% in animal studies. Additionally, we presented results demonstrating the improvement of model match between experiments and forward simulation models by adaptation of GRIN lens optics to a breast phantom study. In particular, 128 GRIN lenses on the fiber bundle face were employed to align the collected excitation and emission light normal to the filter surface in an existing frequency-domain system. As a result of GRIN lens collimation, we reduced the transmission ratio between 10 and 86 % and improved the model match for tomographic reconstruction of one (1 cm3) and two (0.1 cm3) targets in a 1087 cm3 of breast phantom. Ultimately, this work improves the sensitivity of NIR fluorescence imaging by enhancing the rejection of excitation light and shows that the current sensitivity challenges for translating fluorescence-enhanced optical imaging into the clinic can be overcome.