以金奈米粒子與染料結合漸逝波腔體震盪吸收光譜法偵測基質金屬蛋白酶9

Abstract

本研究是利用漸逝波腔體震盪吸收光譜法(evanescent-wave cavity ring-down spectroscopy, EW-CRDS)結合金奈米粒子(gold nanoparticles, Au NPs)與染料rhodamine redTM-X (RRX)作為探針來偵測基質金屬蛋白酶9 (MMP-9)。基質金屬蛋白酶9為人體蛋白酶，其可降解細胞外基質的第四型膠原蛋白。研究發現基質金屬蛋白酶9與腫瘤細胞的轉移有著正相關。罹患癌症的病人其體內有著濃度高於正常人的基質金屬蛋白酶9，因此基質金屬蛋白酶9被視為一種癌症的生物指標。 腔體震盪吸收光譜法的原理是由高反射鏡所構成的腔體來回反射脈衝雷射，並測量由腔體洩漏的能量，將其轉化成腔體震盪時間(ring-down time)。光在腔體內來回反射次數再搭配上腔體的長度，其有效的吸收路徑可達到數公里，可用來偵測微量物質，具有相當高的靈敏性。當腔體內被放置一特殊設計的稜鏡，稜鏡表面因雷射光經全反射所產生的漸逝波可被用來偵測稜鏡表面的光學吸收變化，而此方法被稱作漸逝波腔體震盪吸收光譜法。 本實驗分別使用在表面修飾上鏈親和素(streptavidin)的金奈米粒子與rhodamine redTM-X作為探針。因鏈親和素與生物素(biotin)具有極強的非共價作用力，所以我們在玻片表面修飾上具有生物素的胜肽鏈，並將此玻片放置於產生漸逝波的稜鏡上。而此胜肽鏈對基質金屬蛋白酶9有著非常好的專一性，並且能被其水解成片段。所以當玻片表面的胜肽鏈被基質金屬蛋白酶9水解，玻片表面能吸引修飾上鏈親和素之探針的生物素就會減少，使得吸收改變。漸逝波腔體震盪吸收光譜法可藉此表面之吸收變化，來對基質金屬蛋白酶9進行偵測。

Matrix metallopeptide 9 (MMP-9) belongs to a family of zinc-dependent endopeptidases for degrading the extracellular matrix in human body. MMP-9 has been proven to be a cancer biomarker as MMP-9 is able to help tumor metastasis, and the concentrations of MMP-9 in cancer patients are higher than healthy people. On the other hand, cavity-ring down spectroscopy (CRDS) is an ultra-sensitive method in detecting trace molecules due to long absorption length by multiple reflections inside a cavity composed of highly reflective mirrors. With a customized prism placing inside the cavity, evanescent wave is generated by total internal reflection at the prism surface and able to detect absorption changes. The surface-sensitive method is called evanescent-wave cavity-ring down spectroscopy (EW-CRDS). In this work, EW-CRDS was employed to detect MMP-9 by using two kinds of streptavidin-conjugated probes, gold nanoparticles (AuNPs) and rhodamine redTM-X (RRX) dye, to bind with biotin-labeled peptides on the surface of the prism. This study provides a further sensing method with EW-CRDS.