多光子激发荧光显微成像.pdf

多光子激发荧光显微成像 传统光学显微镜 激光共聚焦显微镜 多光子激发激光扫描显微镜 ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ 早期显微镜 1、光源 2、载物台 3、光学放大系统 ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ 普通显微镜与激光扫描显微镜 不同光源经过样品。（a）普通显微镜的全场照明（b）具有高数值孔径 油镜的激光扫描显微镜，将光聚焦在一点。 ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ 荧光显微镜 Arc Lamp Excitation Filter Excitation Diaphragm Ocular Objective sample Emission Filter 共聚焦显微镜 PMT Objective Emission Filter Emission Pinhole ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ 基本原理 ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ 多光子激发一个非线性过程 两个或多个光子同时激发 n 荧光强度正比于(激光强度) 荧光限制在焦点 ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ ⎯⎯ The Key Laboratory of Biomedical Photonics of Ministry of Education, China ⎯⎯ 多光子与单光子比较 单光子 双光子 * 照明锥体大体积激发 * 激发局限在焦点 * 荧光强度正比于I * 荧光强度正比于In * 光漂白和光损伤较大 * 不需要针孔收集散射光 * 光漂白和光损伤小 * 深度成像 ⎯⎯ The Key Labor