生醫檢測技術常使用抗體-抗原結合技術或是聚合酶連鎖反應，兩者皆需要時間以提高檢體的濃度來進行檢測。其中，核酸物質等需要分離純化之步驟，使檢測過程變得繁複。本研究採用增顯拉曼檢測基板技術，對於目標檢體之結構蛋白進行直接性的檢測，並提高檢測時之靈敏度，屬於免標定之定性檢測與染色標定之相對定量分析。本研究之增顯拉曼檢測基板為「倒多角錐型金奈米壓痕結構陣列」。
奈米壓痕技術可在單一步驟下製作出精密度高之特異性孔洞結構陣列，其物理方式的製程也可以避免檢體受到殘留化學物質的汙染。其中，孔洞結構更可透過極精準之壓深施力與壓痕間距來控制，經由適當控制結構大小之奈米孔洞結構可以誘發表面電漿子侷限化之作用進而產生較大之表面增顯拉曼散射效果。本研究利用羅丹寧6G驗證壓痕結構大小與表面增顯拉曼散射之關係；利用小牛血清蛋白做為標準蛋白來進行免標定之定性檢測與染色標定之相對定量檢測。進一步同時利用具相似分子結構之B型肝炎病毒核心抗原及C型肝炎病毒核心抗原，評估此基板於免標定之定性檢測時之辨別力及再現性。染色標定之相對定量檢測選用具拉曼活性之染劑分子與檢測蛋白結合(如：考瑪斯亮藍G-250)，以建立極低濃度下之相對定量分析。
研究成果顯示：此奈米壓痕結構陣列於免標定之定性檢測時，能夠完整地呈現小牛血清蛋白之指紋特性拉曼圖譜，並能經由拉曼圖譜辨別兩相似分子結構之肝炎病毒核心抗原。染色標定之相對定量檢測濃度可以達到10-7~10-10莫耳濃度，且取檢體濃度之對數與染劑分子之最強特性峰拉曼強度之間呈線性關係。藉由定性及定量檢測之可行性，驗證了此基板的檢測過程可以減少檢測所需時間，且維持其精準度及靈敏度。因此，本研究之檢測基板用具表面增顯拉曼散射；可應用於早期生醫檢測用途。

Bio-medical detection is frequently based on the techniques of antibody-based reactions and polymerase chain reactions. However, these methodologies follow time-consuming incubation steps; to obtain clearly isolated target species requires a complex process. The technique of surface-enhanced Raman scattering (SERS) is competent to provide higher sensitivity, resolution and dynamic analysis with non-labelling qualitative and dye-labelling quantitative analysis. In this study, we introduce “inverted triangular Au nano-indented cavities array” to induce SERS effect upon the substrate.
Nano-indentation technique is a nano-mechanical method which can fabricate particular shape of nano-cavities array in one step, and it is also a physical technique with the advantage of preventing samples from residual chemical contaminations. This method provides an ideal option for the fabrication of nano-structured surface by controlling infinitesimal indented force and tip-to-tip displacements. These nano-cavities arrays would induce the localized surface plasmons (LSPs) effect by properly control the size of cavity structure and lead to significantly enhance Raman signals. Rhodamine 6G (R6G) molecules are used to infer the relationship between the size of cavity structure and SERS effect; bovine serum albumin (BSA) is used as standard protein to estimate un-labelling qualitative ability and dye-labelling quantitative ability; hepatitis B and C virus core antigens are used to estimate the discrimination and reproducibility during the non-labelling qualitative analysis. For dye-labelling quantification, proteins are combined with an existing Raman-active dye (e.g., Coomassie brilliant blue G-250, CBBG-250) to establish a relative quantitative analysis for further detection of low-concentration protein.
These results demonstrated that nano-indented cavities array was competent to clearly characterize the bovine serum albumin with Raman spectrum, even to discriminate the difference of Raman spectra between two varied hepatitis virus core antigens. The distinguishable concentration for dye-lablelling proteins was 10-7~10-10 M within molar concentration, and furthermore there were linear dependent between Raman intensity and the logarithm of the concentration of proteins. Rely on the establishment of qualitative and quantitative techniques, it not only keeps highly sensitivity and accuracy, but also makes up the time-consuming shortage of bio-medical detection. Indeed, the development of SERS-actived substrate in this research could be applied to early bio-medical detection.

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