A Novel Multimodality Nonlinear Microscopy --- Principles and Application

• Main Author: 朱士維
• Other Authors: 孫啟光
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/31783940444643789063

碩士 === 國立臺灣大學 === 光電工程學研究所 === 89 === In this thesis, we report the construction of a nonlinear multimodality microscope based on multi-photon fluorescence, second harmonic generation (SHG), and third harmonic generation (THG). Due to their nonlinear nature, the photodamaging and photobleaching effects can be limited to the vicinity of focus (multi-photon fluorescence) or be avoided at all (SHG and THG). These techniques provide resolution comparable to that of confocal microscopy without aperture and thus raise the sensitivity of the system. Moreover, different physical or chemical characteristics can be picked up by different modalities such as live cell metabolism monitoring by two-photon fluorescence (TPF), tissue structure imaging by SHG, and optical inhomogeneity detection by THG. To demonstrate the usefulness of this new technique, we apply the multimodality microscopy on GaN to determine the relation among the large intrinsic piezoelectric field, the defect-related bandtail state and the optical emission mechanism. GaN-based semiconductor plays a very important role in the light emitting diodes and laser diodes due to their unique emission ability in the blue and the green wavelength regions. In order to obtain more detailed insight into the dominant emission properties of the bulk GaN substrate, we use TPF to show the distribution of bandedge luminescence and defect-related yellow luminescence, which can be an indication to crystal quality. The distribution of bandtail state and the piezoelectric field can be picked up by bandtail state resonant THG and electric field enhanced SHG, respectively. As a result, our microscopy studies indicate the connections between suppressed bandedge PL, increased yellow luminescence, increased bandtail state density, and decreased residue piezoelectric field due to increased defect densities in bulk GaN. As to the InGaN QWs grown on top of the bulk GaN substrate, using 800nm Ti:sapphire laser can avoid the difficulty in interpretation of SHG. By comparing three-photon fluorescence (GaN substrate quality) and SHG (piezoelectric field in QW) images, the result of better substrate quality inducing larger piezoelectric field in QW structure can be obtained. Since one of the major interests of microscopy application is to study the structure and characteristics of biological samples, we then apply the multimodality microscopy to plant cells. With the 1230 nm Cr:forsterite laser, we can acquire not only deeper penetration depth due to the optical transparency in this spectral range, but also the whole nonlinear spectrum in the visible and NIR region, allowing a combination of different imaging modalities including SHG, THG, and multi-photon fluorescence. Our results indicate that while TPF provides information on functional molecules, the intrinsic SHG and THG provide images on specially organized biological sub-cellular nano-structures and biological tissues with optical inhomogeneity respectively. With this technique, we can no only reduce photodamage and photobleaching to a minimum level, but also reduce dye penetration and toxicity problem in live cells.