Intravital fluorescence confocal microscopy has enabled directly in vivo observation of the dynamic 3D cel-lular-level visualization, providing invaluable information impossible to obtain by conventional ex vivo or in vitro experimental methods. In this study, a custom-built real-time laser-scanning confocal microscopy with sub-micron resolution, multicolor, and multi-modal imaging capacity was built and applied to various bio-medical researches.
At first, the effect of THz wave on the biological sample was analyzed due to the increase of interests in a potential biomedical application. Despite the importance of THz wave based applications, yet our knowledge about how THz wave irradiation could affect a live tissue in cellular level are very limited. The immune response in cellular-level was analyzed after pulsed THz wave irradiation generated from the compact FEL in live mouse ear skin in vivo.
In addition, the confocal microscopy system was modified to monitoring ICG, FDA approved biocompatible fluorophore. Many pharmacokinetic studies of ICG on a gross physiological scale were reported due to its clinical value of ICG; however, in vivo cellular-level real-time dynamics has not been well-known. By utilizing the NIR confocal microscopy system, the cellular-resolution ICG and liposomal ICG fluorescence images of the lymph node, the blood vessel in an ear skin, and the liver was successfully visualized in a live anesthetized mouse to investigate pharmacokinetic dynamics in vivo.
Finally, the capability of retinal imaging was achieved by changing the imaging optics of the confocal mi-croscopy platform. Relay imaging optics were carefully optimized for the purpose of retinal imaging in rodent model with an appropriate field of view. Using the system, various pathophysiological studies related with retina was performed, such as monitoring of gene delivery and CNV model, and visualization and tracking of a fluorescent cell.
생체 내 공초점 현미경은 일반적인 ex vivo 또는 in vitro 실험 기법으로는 획득 불가능한 세포들의 역동적인 움직임을 3차원 적으로 관찰 가능한 기법으로, 본 연구에서는 실시간 영상 획득이 가능한 고해상도 레이저 주사 공초점 현미경을 자체 제작 및 변형하여 다양한 의생명 연구에 활용하였다. 우선, 최근 들어 의생명 분야에서 활용 가능성이 높아지고 있는THz 파에 대한 연구로 고출력의 THz 파가 국소적으로 일으키는 염증반응을 앞서 구축한 현미경 기반으로 세포수준에서 분석하였다. 또한, 생체로의 활용에 매우 적합한 FDA 승인된 형광물질인 ICG를 영상화 하기 위한 셋업으로 변형하고, 생체로 도입된 ICG와 리포좀 ICG의 약동학에 대한 연구를 피부혈관, 간, 림프절에서 수행하였다. 마지막으로 다양한 시야각을 가질 수 있도록 망막 영상화 장비를 공초점 현미경 기반으로 구축하고, 망막에서의 유전자 전달, 맥락막 혈관 신생, 형광 세포 및 세포의 동적 움직임을 관찰하였다.
