https://scholars.lib.ntu.edu.tw/handle/123456789/316251
標題: | High-resolution simultaneous three-photon fluorescence and third-harmonic-generation microscopy | 作者: | SHI-WEI CHU CHI-KUANG SUN |
關鍵字: | Laser scanning microscopy; Nonlinear effect; Three-photon excitation | 公開日期: | 2005 | 卷: | 66 | 期: | 4 | 起(迄)頁: | 193-197 | 來源出版物: | Microscopy Research and Technique | 摘要: | In recent years, nonlinear laser scanning microscopy has gained much attention due to its unique ability of deep optical sectioning. Based on our previous studies, a 1,200-1,300-nm femtosecond laser can provide superior penetration capability with minimized photodamage possibility. However, with the longer wavelength excitation, three-photon-fluorescence (3PF) would be necessary for efficient use of intrinsic and extrinsic visible fluorophores. The three-photon process can provide much better spatial resolution than two-photon-fluorescence due to the cubic power dependency. On the other hand, third-harmonic-generation (THG), another intrinsic three-photon process, is interface-sensitive and can be used as a general structural imaging modality to show the exact location of cellular membranes. The virtual-transition characteristic of THG prevents any excess energy from releasing in bio-tissues and, thus, THG acts as a truly noninvasive imaging tool. Here we demonstrated the first combined 3PF and THG microscopy, which can provide three-dimensional high-resolution images with both functional molecule specificity and submicrometer structural mapping capability. The simultaneously acquired 3PF and THG images based on a 1,230-nm Cr:forsterite femtosecond laser are shown with a Hoechst-labeled hepatic cell sample. Strong 3PF around 450 nm from DNA-bounded Hoechst-33258 can be observed inside each nucleus while THG reveals the location of plasma membranes and other membrane-based organelles such as mitochondria. Considering that the maximum-allowable laser power in common nonlinear laser microscopy is less than 10 mW at 800 nm, it is remarkable that even with a 100-mW 1,230-nm incident power, there is no observable photo damage on the cells, demonstrating the non-invasiveness of this novel microscopy technique. ? 2005 Wiley-Liss, Inc. |
URI: | http://www.scopus.com/inward/record.url?eid=2-s2.0-20544436237&partnerID=MN8TOARS http://scholars.lib.ntu.edu.tw/handle/123456789/316251 |
DOI: | 10.1002/jemt.20160 | SDG/關鍵字: | 4 [5 (4 methyl 1 piperazinyl)[2,5' bi 1h benzimidazol] 2' yl]phenol; chromium; DNA; article; cancer cell culture; carcinoma cell; cell membrane; cell strain HepG2; chromium forsterite femtosecond laser; energy; fluorescence microscope; fluorescence microscopy; human; human cell; image quality; image reconstruction; imaging and display; laser; laser microscopy; non invasive measurement; photon; priority journal; spectral sensitivity; third harmonic generation microscopy; three dimensional imaging; three photon fluorescence microscopy; Bisbenzimide; Carcinoma, Hepatocellular; Cell Line, Tumor; Humans; Image Enhancement; Liver Neoplasms; Microscopy, Confocal; Microscopy, Fluorescence, Multiphoton |
顯示於: | 電機工程學系 |
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