微米與奈米級雷射顯微摘取用轉移膜之研究

The Study of Transfer Film for Micron and Nano Laser Capture Microdissection System

沈傳宇(Chuan-Yu Shen)

指導教授 ： 陳建銘

國立臺北科技大學/機電學院/光電工程系所/碩士(2006年)

摘要

本研究主要為微米與奈米級的雷射顯微摘取(Laser Capture Microdissection, LCM)用之轉移膜開發。顯微雷射摘取的原理係將鍍有轉移膜面的載具置於組織切片上方，雷射光經由顯微物鏡穿透載具，聚焦於轉移膜，該處轉移膜吸收光能轉為熱能，產生熔化並黏著目標細胞，當載具從切片上方移除，即可分離出目標細胞。此轉移膜的熔點為88℃，對於生物細胞易造成破壞，且雷射光穿透壓克力載具易產生由吸收及反射所造成的功率損耗等缺點，因此本研究研發新的EVA材料及新的載具，以作為傳統型及近場光纖探針型雷射顯微摘取系統之使用。
實驗使用國產之乙烯醋酸乙酯(Ethylene Vinyl Acetate, EVA)為主要材料，熔點為65℃，透過加入近紅外光吸收染料以提升吸收性，經由調配EVA溶液、基板製備、旋轉塗佈等製程，以完成不同濃度、厚度的各式轉移膜，另外亦設計中空載具及薄膜鍍於載具的製程，藉以減低功率損耗的缺點。
轉移膜的評價是使用光譜分析、原子力顯微鏡(AFM)、掃瞄式電子顯微鏡(SEM)等，找出較佳之吸收熔化情形，以決定製程參數，此參數有近紅外光染料濃度、薄膜表面平坦度等，此結果並與傳統型轉移膜比較。最後，透過近場光纖探針型雷射顯微摘取系統的使用，來發現不同膜厚及熔化圓點大小的關係，當膜厚為30μm、40μm、50μm下分別可產生225nm、320nm、500nm的熔化範圍，最後完成20奈米之單層金粒子的摘取，摘取的最小直徑可達200nm。本論文成功完成了應用於微米級與奈米級的雷射顯微摘取用之轉移膜。

關鍵字

轉移膜 ； 乙烯醋酸乙酯 ； 旋轉塗佈

並列摘要

The purpose of the study is to provide the innovation of transfer film for Micron and Nano Laser Capture Microdissection (LCM) system. The principle of LCM comes from placing a transfer film on the top of dissected tissue. As laser beam pierce on the transfer film, the penetrated spot then converts and absorbs laser energy into heat to adhere the target cell after melting. Then the target cell will be identified after transfer film is removed from the dissected tissue. In order to make it flat and convenient to use, transfer film is coated on the bottom of a cap. But the cap may cause negative impacts such as power loss after the absorption and reflection of laser beam. Therefore, this study is aiming to find the new type EVA cap for the usage of traditional and probe type Laser Capture Microdissection.
The melting point of the transfer film is 88℃, which may damage cells easily.
We use local-made EVA, which melting point is 65℃, as major material and add near-infrared absorbing dyes to enhance the laser energy absorption. By combining different concentration of EVA solution, substrate preparation, and spin-coating etc., we made various transfer films with different concentration and thickness.
In this study, we designed a hollow cap, which made the fiber probe exert close melting, and the optimal conditions of making the thermoplastic film adhere on the cap were also studied.

並列關鍵字

Laser Capture Microdissection(LCM)

參考文獻

[4] W. M. Ruge, W. Bielser, E. Remy, F. Hillenkamp, R. Nitsche, R. Unsold, "The laser in the Lowry technique for microdissection of freeze-dried tissue slices," Histochemical journal, 8, 1976, pp.387-401.

[7] C. F. Yen, J. A. Lee, C. M. Chen, "New Construct of Laser Capture Microdissection(LCM) Using Near-Field Optical Tips," The 10th International Microoptics Conference(MOC’4), Germany, 2004.

[8] M. B. Thomas, F. H. David,"Laser capture microdissection analysis vessel," U.S. pat. 5859699, 1997.

[10] M. B. Thomas, F. H. David, J. S. Toeppen," Broadband absorbing film for laser capture microdissection," U.S. pat. 6495195, 2003.

[11] M. Rubin," Use of laser capture microdissection, cDNA microarrays, and tissue microarrays in advancing our understanding of prostate cancer," Journal of Pathology, 195, 2001, pp.80-86.