A modular fiber optic system for intramural functional fluorescence measurement

摘要

Fluorescence imaging techniques have been central to much biomedical science research over the past two decades. In particular, functional imaging has provided important new information about processes that occur at cellular and sub-cellular levels. With this approach, living tissues are stained with dyes whose emission is modulated by changes in the environment to which the dye is exposed. The fluorescence imaging systems used within this context typically incorporate relatively complex free space optical assemblies and a stable platform is necessary to maintain appropriate alignment of their components. Because of the poor efficiency of these systems, it is necessary to use powerful light sources and sensitive photo-detectors. We have developed a novel fluorescence imaging system in which free-space optics are replaced by optical fibers, passive optical splitters and associated components. Solid state lasers are used as the excitation light source. A variety of detection systems have been utilized including a spectrometer. The feasibility of the approach has been established using a rat heart preparation stained with the membrane potential-sensitive dye, di-4-ANEPPS. Detailed emission spectra for this dye, at different levels of resting membrane potential, are presented here for 532 nm and 488 nm excitation. Cardiac action potentials obtained with the modular fiber optic system correspond closely to intracellular potentials acquired at adjacent sites in the isolated rat heart preparation. Our modular fiber optic system is cheaper, more efficient, more flexible and more robust than conventional fluorescence imaging systems. Using a high-speed spectrometer for photodetection, it is possible to implement the signal processing required for multi-line or ratiometric imaging in software, which further enhances the efficiency and flexibility of the system. We believe that this approach has wide potential applications for biomedical fluorescence imaging.