基於雙折射晶體之相位式全像式表面電漿子共振感測器

Abstract

根據許多文獻指出，相位式表面電漿子共振感測器比強度式更加靈敏，對折射率之解析度至少高一個數量級。基於相位檢測在靈敏度上的優勢與棲列式生物分子感測的需求，我們提出一個新型偏振干涉的相位式影像式表面電漿子共振感測器。 本論文提出基於雙折射晶體之相位式全像式表面電漿子共振感測器，本技術結合垂直共振腔面射雷射本身的特性及雙折射晶體之特殊晶體結構達到有效的相位調變與干涉。主要是利用雙折射晶體在兩軸向上不同的折射率，對P 和 S 波造成相位差，進而在極化片的幫助下使得兩軸向的資訊投影到同一平面產生干涉，配合團隊先前已獲有專利的泛用式鎖相放大器，擷取其相位及強度資訊。 為了證明本研究所提出之系統架構的可行性，我們構建了一個原型系統來進行驗證。在此我們透過雷射波長-電流調變係數(S)及特定之電壓調變等特殊條件下，達到特定的相位深度(∆ϕ=3.8317)並解調出相位資訊。利用影像式的優點，我們能篩選干涉較好的區域進行量測，同時提升訊雜比。在此架構下，本系統可在雜訊0.0004 rad的層級下達到7.5∙10^-7 RIU之最低偵測極限。而根據模擬我們推估，電漿層膜厚為49 nm下可以進一步提升靈敏度。最後，透過K-ras基因突變檢測，本研究驗證了其在生物感測方面之應用潛力。在本系統下，我們能量測到50 nM濃度之K-ras基因突變樣本。此外，由於通過多陣列測量K-ras之單點突變的影響，我們初步驗證了本系統對單核甘酸多態(Single Nucleotide Polymorphism , SNP）的分辨能力。最後，利用動力學分析以確認我們最終結果之可靠性。

Surface Plasmon Resonance (SPR) sensor with optical phase interrogation has been well known to exceed the performance of conventional SPR by at least one order of magnitude. To take the advantage of phase interrogation SPR, we propose herein a phase-sensitive imaging SPR system based on a novel interferometric scheme. The basis of the proposed technology is the use of Vertical Cavity Surface Emitting Laser (VCSEL) diode which provides cost-effective phase modulator to discriminate amplitude and phase. VCSEL is driven in current to create minute wavelength modulation such that sufficient phase modulation can be achieved (∆ϕ=3.8317). The interference is generated with the help of a YVO4 birefringent crystal, which leads to phase difference between S and P polarization. After the measuring beam passing the SPR sensor chip, the S and P polarization of the beam are then projected back into a single polarization direction, thereby generating polarimetry type interference. Moreover, since the amplitude is modulated at same frequency with phase, the phase retrieval a non-trivial task. We have therefore adopted a previously published Generalized Lock-In Amplifier (G-LIA) to acquire phase information. To demonstrate the efficacy of the proposed system, we built a prototype system to carry out proof-of-concept. First by estimating wavelength-to-current sensitivity factor (S) value, which is intrinsic nature of individual VCSEL, we determine a specific current modulation that is needed to achieve ∆ϕ=3.8317. We then demonstrate the feasibility to the proposed phase sensitive SPR system through fringe image and SPR image at Krestchmann angle. To accurately distinguish signal of DNA measurement, thermal drift issue is subtracted by adding a reference arm. Then, with glucose solution for sensitivity calibration, we demonstrate a Limited-of Detection (LOD) of 7.5∙10^-7 RIU where noise is estimated to be around 0.0004 rad. From our data and simulations, we estimate that sensitivity can largely improve at around film thickness 49 nm. Finally, a result of K-ras mutation sensing down to 50nM is presented. Also, with the effect on mismatch K-ras DNA measured through multi-array, we proof that our system is capable to distinguish Single Nucleotide Polymorphism (SNP). Concentration calibration together with kinetics analysis is presented to confirm the reliability of our final results.