Fluorescence microendoscopy imaging based on GRIN lenses with one- and two-photon excitation modes

Abstract

With the rapid development of life sciences, there is an increasing demand for intravital fluorescence imaging of small animals. However, large dimensions and limited working distances of objective lenses in traditional fluorescence microscopes have limited their imaging applications mostly to superficial tissues. To overcome these disadvantages, researchers have developed the graded-index (GRIN) probes with small diameters for imaging internal organs of small animals in a minimally invasive fashion. However, dynamic imaging based on GRIN lens has not been studied extensively. Here, this paper presented a fluorescence endoscopic imaging system based on GRIN lenses using one-photon and two-photon excitation. GRIN lenses with 1.15 mm diameter and 7.65 mm length were used in the system. The images were acquired by a compact laser scanning imaging system with a resonant galvo-mirror system to scan the laser beam and a photomultiplier tube (PMT) to detect fluorescence signals. Experimental results showed that this system using two-photon excitation could implement dynamic fluorescence microendoscopic imaging and monitor the movement of blood flow beneath the skin in anesthetized mice while producing images with higher contrast and signal to noise ratio (SNR) than those using one photon excitation. It would be a useful tool for studying biological processes of small animals or plants in vivo.