表面電漿共振(Surface Plasmon Resonance，SPR)為可利用檢測光學直接偵測晶片表面上因生物反應造成折射率微量變化來達到免標定之生醫光電技術。SPR 技術具有靈敏度遠高於其他檢測方法且可排除標定的繁複過程，並且能即時地分析生物分子之間的交互作用(biomolecular interaction analysis，BIA)，而大大地縮短檢測的時間並提供了更多且精確的生物資訊。近年來，已被廣泛地應用在很多生物分子診斷的領域上，諸如receptor-ligand 或抗原(antigen)與抗體(antibody)之交互作用、蛋白質分子的非特定吸附、薄膜與蛋白質間交互反應或去氧核醣核酸分子雜交(DNAhybridization)等。 本實驗室在本校機械系陳顯禎教授之協助下，自行研發的表面電漿共振系統已成功應用於各式生物分子診斷等領域，如建立完整的核酸雜交動力學研究程序，並實際檢測寡去氧核糖核酸序列雜交試驗。使用此系統於檢測、研究分子間的交互反應動力學機制可提供一套非常準確、即時的資訊。 本研究是利用表面電漿共振儀，配合DNA 分子結構預測軟體(Oligo)所計算之DNA 二級結構，來量測不同二級結構之單股DNA 於不同溫度下雜交的行為。藉此得知處於不同雜交環境以及不同的DNA 分子結構下雜交反應之動力數據，再配合質傳限制模式(Mass transport limitation model)以動力學的角度進行數據分析，並使用Van’t Hoff 方程式在DNA 雜交反應之熱力學數據對DNA 雜交反應的機制作較詳細的討論。 This study utilizes a surface plasmon resonance (SPR) biosensor and a theoretical secondary structure calculation program(Oligo) to investigate the influence of secondary structures of ssDNA on the DNA hybridization. It is found that the SPR angular shifts associated with the three pairs of 60mer oligo-nucleotides with prominent secondary structures are lower than those observed for the two pairs of oligonucleotides with no obvious secondary structures. It is also determined that increasing the DNA hybridization temperature from 35 oC to 45 oC reduces secondary structure effects on hybridization. On the hybridization with mixture target oligo-nucleotides, the SPR results demonstrate that presence of oligo-nucleotides mixture due to non-specific interactions between non-complementary probes and targets interfere the extent of kinetics of hybridization significantly. In this investigation, thermodynamics information was obtained by Van’t Hoff equation and combining with the kinetics data from SPR, the hybridization mechanism were proposed and discussed.
