應用色胺酸合成金奈米粒子於比色法檢測尿液血清白蛋白

Abstract

本實驗中，我們成功開發金奈米比色法探針檢測人類血清白蛋白。我們添加氫氧化鈉於色胺酸與四氯金酸混合溶液中合成色胺酸金奈米粒子。藉由人類血清白蛋白與色胺酸之間的配位結合，人類血清白蛋白會因為與金奈米粒子表面的色胺酸作用，保護金奈米粒子以避免在5×磷酸鹽緩衝溶液中發生鹽類誘導聚集的現象，溶液仍呈紅色。但沒有人類血清白蛋白保護的金奈米粒子，色胺酸金奈米粒子在5×磷酸鹽緩衝溶液中會發生鹽類誘導聚集，使溶液呈藍紫色。為了確認人類血清白蛋白與色胺酸的配位結合，我們添加界面活性劑將蛋白質變性或添加競爭配位結合的藥物與人類血清白蛋白作用。實驗結果說明這些都會影響人類血清白蛋白與金奈米粒子表面色胺酸的配位結合，使得色胺酸金奈米粒子於高鹽環境下發生鹽類誘導聚集。此外，相較於其他蛋白質，只有血清白蛋白在最佳化5×磷酸鹽緩衝溶液(pH 7.4)的條件下能保護金奈米粒子。隨著不同濃度的人類血清白蛋白，用肉眼便能觀測到100 nM人類血清白蛋白能造成的色胺酸金奈米粒子顏色變化。藉由紫外/可見光分光光譜儀，可以檢測人類血清白蛋白的最低可偵測濃度約為60 nM。在尿液樣品應用方面，其最低可偵測濃度為120 nM。因此色胺酸金奈米粒子可以做為高選擇性與高靈敏度的比色探針檢測人類血清白蛋白。

A gold nanoparticle-based colorimetric probe for the detection of human serum albumin (HSA) was developed in this study. We synthesized the tryptophan-stabilized gold nanoparticles (Trp-AuNPs) by mixing tryptophan with tetrachloroauric acid at the alkaline solution. Due to the ligand binding between HSA and tryptophan, HSA would interact with the tryptophan on gold nanoparticles surface and prevent the Trp-AuNPs from the salt-induced aggregation. The Trp-AuNPs were red in color and exhibited absorption at 530 nm. Without HSA, the salt-induced aggregation of Trp-AuNPs would occur in 5× phosphate buffer saline (PBS) and result in the appearance of a new absorption at 700 nm. To confirm the ligand binding between HSA and tryptophan, we used the surfactants to denature HSA or added a competitive ligand binding drug to interact with HSA. The results showed that both of them would influence the interaction between HSA and tryptophan and cause the Trp-AuNPs to aggregate. Compared with several proteins, only the serum albumin could prevent the Trp-AuNPs from salt-induced aggregation under the optimized condition (5× PBS, pH 7.4). The concentration of HSA can be visualized about 100 nM by the naked eyes through the color change of Trp-AuNPs and determined by UV-vis spectroscopy with the lowest detectable concentration of 60 nM. Therefore, Trp-AuNPs can be employed for highly selective and sensitive colorimetric detection of HSA.