本計畫主要以發展表基因體學（epigenetics）之影像相關分析技術為主要目標。表基因體學為目前熱門之研究主題，研究表基因體學主要以傳統聚合??鎖反應、DNA 定序、微陣?晶片等商業化產品為主要分析工具。本計畫書之緣起是接續上??計畫：以毛細管電泳暨?射誘發螢光為基礎開發癌症相關生物標的之新穎檢測技術（NSC 98-2113-M-029 -008 -MY2），以毛細管電泳暨?射誘發螢光為分析工具偵測到被病毒感染之鼻?癌細胞內的微小核醣核酸（microRNA, miRNA）。本計畫初期則以全內反射螢光（total internal reflection fluorescence, TIRF）技術為基礎發展高?敏?影像技術直接於細胞或病人組織內觀察、偵測 miRNA。從101 ??起，將緊接著以TIRF 開發DNA 甲基化分析技術偵測癌細胞之甲基化程?與?質性，期望可以提供生物醫學相關研究人員一個新穎、可信賴之DNA 甲基化分析方法。 In this proposal, we design an ultra-sensitive imaging system for epigenetics based on total internal reflection fluorescence (TIRF). A high power laser beam is split by beam splitter. The produced two laser beams separately focus on right-angle prism and high-NA objective, thereby simultaneously introducing two evanescent waves on an inverted microscope. The evanescent wave-induced fluorescence will then generate on the surface of two cover slide after fluorescent, specific bio-recognition molecule. This method may allow us to detect the epigenetic-related biomarkers, such as low-abundance microRNAs within cell or tissue. In the 2nd year, we develop a TIRF-based imaging system for the analysis of DNA methylation. Our previous results demonstrated that DNA methylation distributed differently in various cancer cell. Thus, we assume that heterogeneity of DNA methylation play an important role in cancer biology. To achieve this goal, the combination of TIRF microscope and specific antibody recognition is utilized to explore the state of DNA methylation in single DNA molecule.
