探討溫度對綠螢光蛋白光學特性之影響

Study on the Effects of Temperature on the Optical Properties of Green Fluorescent Protein

蔡宜靜(I-Ching Tsai)

指導教授 ： 許怡仁

中原大學/理學院/物理研究所/碩士(2010年)

https://doi.org/10.6840/cycu201000570

摘要

由於綠螢光蛋白具有高穩定性與高量子效率的優勢，近來已成為研究活細胞和蛋白質的重要螢光標定。本研究觀察綠螢光蛋白在不同溫度之下的螢光光譜變化，並探討螢光蛋白在長時間的脈衝雷射激發之下所發生的光漂白現象。我們發現綠螢光蛋白的螢光光譜存在來自五組不同能帶間躍遷的訊號。本研究於－30 ℃至80 ℃的溫度範圍之內量測綠螢光蛋白的螢光光譜，發現螢光的強度與光譜分布皆隨溫度而有顯著的變化。此外，推測綠螢光蛋白的電子在不同能帶上的分布亦隨著溫度而產生了變化。在光漂白的研究上，發現綠螢光蛋白在脈衝雷射的激發之下，螢光強度會隨時間呈現雙指數衰減的行為，且此行為亦隨溫度的變化而有所差異。

關鍵字

溫度相關 ； 綠螢光蛋白 ； 螢光光譜 ； 光漂白

並列摘要

Enhanced green fluorescent protein (EGFP) possesses the advantages of high stability and high luminescence. Therefore, it has become an important fluorescent label in the researches of cells and proteins. In this research, the dependence of the fluorescent spectra of EGFP on temperature and the effect of photobleaching under the excitation of laser pulses were investigated. We found that the fluorescent spectrum of EGFP consists of five components that might come from the transitions between different electronic energy states. Furthermore, it was observed that the intensity of the fluorescence and the distribution of the spectrum are strongly dependent on the temperature in the range from –30 °C to 80 °C. It is speculated that the distribution of electrons in the excited states changes with temperature. In the study of photobleaching, we found that the intensity of EGFP has the behavior of double exponential decay with time. The effect of photobleaching of EGFP is also dependent on temperature.

並列關鍵字

fluorescent spectrum ； green fluorescent protein ； photobleaching ； temperature dependence

參考文獻

[30] 曾盈智（民國九十二年），《紅螢光蛋白及綠螢光蛋白生化特性的研究》，嘉義大學生物科技研究所。

[6] O. Nakamura and S. Kawata, “Three-dimensional transfer-function analysis of the tomographic capability of a confocal fluorescence microscope.” J. Opt. Soc. Am. A7(3), 522-526 (1990).

[7] K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope.” Rev. Sci. Instrum. 78(2), 023705 (2007).

[8] N. Bozinovic, C. Ventalon, Tim Ford, and Jerome Mertz, “Fluorescence endomicroscopy with structured illumination.” Opt. Express 16(11), 8016-8025 (2008).

[9] O. Shimomura , Frank H. Johnson , Yo Saiga, “Extraction, Purification and Properties of Aequorin, a Bioluminescent Protein from the Luminous Hydromedusan, Aequorea.” J. Cell. Comp. Physiol. 59, 223-29 (1962).