Development of aggregation induced emission-boosted adaptive optical (AIE-AO) confocal microscopy for enhanced bioimaging

Abstract

Confocal microscope is widely used in bioimaging because of its optical sectioning capability, high resolution, and low cost. However, as the imaging depth increases, aberrations induced by the specimen or the optical system significantly degrade the imaging resolution and contrast, which hinders the application of confocal microscopy in deep-tissue imaging. My M.Phil. research focuses on studying the nature of different types of aberrations in thick and highly scattered samples and aims to correct the aberrations using indirect adaptive optics (AO) methods to achieve optical imaging through severely scattered media. Aggregation-induced emission (AIE) material is used as a guidestar to further enhance the imaging depth and contrast. In this thesis, the experimental results have demonstrated that our self-developed adaptive optical confocal microscope performs well in various bioimaging applications with acceptable resolutions. The proposed aberration correction method with AIE material has also been proved to be effective in imaging deep in highly scattering media. The thesis is organized as follows. In the first part, the mechanisms of confocal microscopy, adaptive optics, and aggregation-induced emission material are introduced. The influence of aberrations on confocal imaging system and the limitations of present fluorophores are also discussed. To overcome the current challenges, an aggregation-induced emission-boosted adaptive optical (AIE-AO) confocal microscopy system is proposed to achieve high-resolution, high-contrast, and deep-penetration optical imaging in scattering samples. The second part of this thesis focuses on the adaptive optics algorithm, the design of the confocal microscope, and the control system. For the hardware, the key components and structures of the imaging system are introduced in detail; for the software, the process of three-dimensional imaging, system calibration, and wavefront modulation are explained. In the third part, the thesis demonstrates the performance of the AIE-AO confocal microscopy system. Fluorescent beads, scattering phantoms, and mouse brain slice have been used as samples to show the improvements in image quality with adaptive optics. After that, the emission properties of the AIE material in highly scattered phantoms are compared with a commonly used fluorophore, Nile Red. In conclusion, the study has demonstrated that the AIE-AO confocal microscopy system can significantly improve the imaging signal intensity, resolution, contrast, and penetration depth in scattering samples. The signal intensity has been improved around three to four times, meanwhile the lateral resolution also improved by 60% at around 200 μm depth with AO-AIE boosted. While further improvements are needed, it is believed that the homebuilt system will have sound application potential in bioimaging and generate exciting future explorations.