以表面電漿波共振與奈米金標記技術探討硫醇與牛血清蛋白之結合反應   免費試閱

Binding Reaction Analysis between Thiol and Bovine Serum Albumin by Surface Plasmon Resonance and Au Nanoparticle Labeling Technologies

由於薄膜式表面電漿波共振技術檢測無法檢測微小分子之結合反應訊號，因此，本研究使用奈米金標記小分子之技術來增強反應訊號，此處主要進行有奈米金標記之11 個碳之硫醇分子的反應動力學分析，由於硫醇分子尾端可以是羧基或是可由1-（3- 二甲氨基丙基）-3- 乙基碳二亞胺（EPC）及N- 羥基丁二醯亞胺（NHS）活化轉變成NHS 酯，在注入流道後可以靜電吸附或胜肽鍵結方式與固定於感測器表面之牛血清蛋白之胺基進行結合，其中，胜肽鍵結所獲得之飽和表面電漿波共振角位移可達0.8° 而靜電吸附可達0.365°，其主要是具NHS 酯之硫醇修飾之奈米金未帶有表面電荷，因此可以較密集地鍵結於感測器表面；相反地，中性pH 值下具羧基之硫醇修飾之奈米金帶有負表面電荷會因同性相斥而無法密集地結合於感測器表面，本研究也探討這些樣品互為流動相或固定相所產生之結合作用，結果顯示，流動相為牛血清蛋白與固定相為具NHS 酯之金奈米子之結合速率常數高達56 × 103sec-1M-1，而若與固定相為具NHS 酯之奈米金膜之結合速率常數則為9 × 103sec-1M-1。另外，具羧基之金奈米子以靜電吸附到固定相過程中皆會有脫離現象產生。

For thin film type surface plasmon resonance detection technology, it is hard to measure the small molecule binding reaction signal. Au-nanoparticle labeled small molecule technology is used to enhance the response signal in this paper. We mainly focused on the reaction dynamics analysis of the Au-nanoparticle labeled thiol molecules having 11 carbon atoms. Because the end of the thiol molecules can be carboxyl group or NHS-ester after EDC and NHS activation, thiol molecules injected into a flow channel can form electrostatic absorption or peptide bonding to the bovine serum albumin immobilized on the sensor surface. Saturated surface plasma resonance angle shift can be as large as 0.8° for the peptide bonding and 0.365° for the electrostatic adsorption. For peptide bonding thiol-modified Au nanoparticles with electrically neutral NHS-ester can densely arrange on the sensor surface, on the contrary, the thiol-modified gold nanoparticles with electrically negative carboxyl group cannot due to the repulsive electrostatic force between nanoparticles. Here we also discuss the effect for changing the conditions of the samples among mobile and immobile phases. The results show the association rate constant for the mobile bovine serum albumin to the immobilized Au-nanoparticles with NHS-ester has a value as high as 56 × 103sec-1M-1, however, for the gold film with NHS-ester is 9 × 103sec-1M-1. Another results show the electrostatic adsorption between the carboxyl modified Au nanoparticles and other immobile samples are always disrupted.