双模态全视场光学相干层析技术的角膜缘高分辨率成像

High-resolution imaging of limbus tissue with dualmode full-field optical coherence tomography

目的:开发细胞级高分辨率、结构与功能一体化的双模态全视场光学相干层析系统(full-field optical coherencetomography,FFOCT),实现角膜缘组织的双模态FFOCT成像。方法:基于Linnik干涉成像原理,利用高数值孔径显微物镜(NA=0.8)及高速平面互补金属氧化物半导体(complementary metal oxide semiconductor,CMOS)相机,设计搭建高分辨率的组织静态结构和内源动态功能成像一体化双模态FFOCT系统;构建基于四相位调制结构影像提取及时域干涉信号动态频谱分析的功能影像重建算法;对人供体角膜缘组织开展各深度层的双模式FFOCT成像有效性验证。结果:搭建的双模态FFOCT成像系统可实现横向分辨率0.5μm,轴向分辨率1.7μm,成像视野320μm×320μm,相机采集速度100 Hz。系统实现角膜缘组织无外源标记情况下的细胞级分辨率三维结构和内源功能成像,FFOCT静态结构影像清晰显示角膜缘上皮、Vogt栅栏、隐窝、基质、血管及淋巴管等结构,FFOCT动态功能影像突出显示了代谢活跃细胞(角膜缘上皮细胞、免疫细胞等)。结论:双模态FFOCT高分辨率成像系统可提供角膜缘微观结构和活细胞无标记内源功能可视化信息,将为角膜缘疾病的研究及临床诊疗提供全新的成像分析技术。

Objective:To develop a cellular-level,high-resolution,integrated dual-modal full-field optical coherence tomography(FFOCT)system capable of simultaneously imaging the structure and function of limbus tissue.Methods:Utilizing the Linnik interference imaging principle,a high-resolution dual-modal FFOCT system was designed and constructed using a high numerical aperture(NA=0.8)microscope objective and a high-speed flat CMOS camera.A functional imaging reconstruction algorithm based on four-phase modulation structure image extraction and dynamic frequency spectrum analysis of temporal interference signals was developed.The effectiveness of dual-mode FFOCT imaging at various depth layers of human corneal limbal tissue was validated.Results:The constructed dual-modal FFOCT imaging system achieved lateral resolution of 0.5μm,axial resolution of 1.7μm,imaging field of view of 320μm×320μm,and camera acquisition speed of 100 Hz.The system enabled cellular-level resolution three-dimensional structural and intrinsic functional imaging of corneal limbal tissue without exogenous labeling.Static structural FFOCT images clearly displayed limbal epithelium,palisades of Vogt,crypts,stroma,blood vessels,and lymphatic vessels,while dynamic functional FFOCT images highlighted metabolically active cells(limbal epithelial cells,immune cells,etc.).Conclusion:The dualmodal FFOCT high-resolution imaging system provides visualization of corneal limbal microstructural and live cell intrinsic functional information without labeling,offering a novel imaging analysis technique for research and clinical diagnosis and treatment of limbal diseases.