矽光子晶體增強螢光在蛋白質陣列晶片的應用

Abstract

本研究計畫的目的是要發展矽光子晶體基板來增強螢光並改善檢測極限，並應用於蛋白質陣列晶 片。預期達成的目標包括：1. 根據嚴格耦合波分析法來最佳化光子晶體的結構，2. 建立相位光罩 微影技術來製作大面積且均勻的矽光子晶體，3. 評估光子晶體和一般玻璃的螢光增強的效應，4. 使 用微陣列三明治酵素連結免疫吸附法來檢測腫瘤壞死因子及比較光子晶體和一般玻璃的檢測敏感 度。 本計畫的光子晶體在功能上為一個光共振器，是一個包含高折射率介電質覆蓋在低折射率的光柵 結構。本結構在特定組合的入射波長及角度將存在兩個共振模態。其中一個共振模態將會和激發雷 射光波長產生光譜匹配，來增強近場電場強度以達到增強激發(enhanced excitation)。另一個共振 模態將會和螢光發射波長產生光譜匹配，使螢光能更有效率地被導向偵測儀器來達到增強萃取 (enhanced extraction)。有效的結合這兩種機制，可使螢光檢測的方法具備更佳的敏感度。 除了微陣列晶片的應用，矽光子晶體螢光增強技術可以輕易地整合到其它螢光分析、檢測應用的 基板，並應用到各種不同的生物研究領域，例如基因表現分析(可檢測更低濃度的基因)、蛋白質分 子檢測(可檢測更低濃度的腫瘤標記來及早篩選癌症)、細胞顯微技術(可觀測更低濃度螢光標記的細 胞內部組成)。

The goal of this project is to develop a Si photonic crystal (PC) enhanced fluorescence platform to improve the detection limit for microarray applications. To do so, two different PC designs are proposed. Our aims are to: 1. optimize the PC geometries based on Rigorous Coupled Wave Analysis (RCWA), 2. establish phase mask lithography capability for large-area and consistent fabrication of Si PCs, 3. evaluate the fluorescence enhancement among two different designs and regular glass slides and 4. demonstrate the improved sensitivity for detection of tumor neccrosis factor-alpha (TNF-α) using a sandwich enzyme-linked immunosorbent assay (ELISA) in microarray format. The PCs used in this work are subwavelength nanostructures composed of a periodically modulated low refractive index layer that is coated with a high refractive index dielectric and are designed to perform as an optical resonator. The device will exhibit two distinct resonances at a specific combination of wavelength and angle of incidence. One resonance will spectrally match with the excitation laser wavelength for near field enhancement to have “enhanced excitation.” Meanwhile, another resonance will spectrally overlap with the fluorophores’ emission to more efficiently direct the emitted photons towards the detection instrument to obtain “enhanced extraction.” Simultaneous implementation of two mechanisms can maximize the fluorescence output to achieve better sensitivity for fluorescence-based detection. With successful development, besides microarray applications, Si PC enhanced fluorescence platforms can be incorporated into many surfaces that support fluorescent assays or measurements. Depending on the design of the detection instrument, researchers will be able to take advantage of enhanced extraction, enhanced excitation or both effects simultaneously. The developed capability will have broad implications for gene expression analysis (the ability to observe presence of genes that are expressed at lower concentrations than currently observable), molecular diagnostics (the ability to detect presence of disease biomarkers at lower concentrations), cell microscopy (the ability to observe cells labeled with low efficiency fluorescent proteins), among many other fields.