簡易膠體電泳法用於奈米粒子穩定性檢測：白蛋白誘導不穩定聚乙二醇化金奈米粒子調理素作用之快速感測

General Gel Electrophoretic Protocol for Monitoring Nanoparticles Instability: Albumin Induce the Rapid Sensing of Opsonization onto the Unstable PEGylated Gold Nanoparticles

王銘翊(Ming-Yi Wang)

指導教授 ： 林政鞍

中原大學/工學院/生物醫學工程研究所/碩士(2018年)

https://doi.org/10.6840/cycu201800397

摘要

本研究提供一個嶄新的膠體電泳策略，簡易且快速的進行奈米粒子穩定性檢測。此檢測方法能應用於多種尺寸之聚乙二醇奈米粒子，能明確分辨各奈米粒子之聚乙二醇配體密度、配體金硫鍵結穩定性及環境老化測試，檢測技術成本低廉但檢測成果卻相對有效。實驗透過半胱胺酸配合牛血清白蛋白對聚乙二醇奈米粒子進行調理素化，並改變奈米粒子表面電性，依照電性及電泳速率差異在膠體電泳上進行分離。研究成功利用牛血清白蛋白作為調理素對金奈米粒子表面聚乙二醇密度進行檢測，同時配合脫附劑半胱胺酸進行金奈米粒子的強制調理素化，成功且明確的表現出了聚乙二醇金奈米粒子於脫附劑環境中的不穩定性，實驗同時證明了穩定性檢測可透過熱處理進行反應加速，更進一步縮短奈米粒子調理素化檢測的時間。此外透過膠體電泳與紫外可見光吸收儀、表面電位分析儀的結果比較發現，膠體電泳檢測相較於前者能更明確快速的表現出穩定性差異。而在熱處理實驗的過程中另外發現金奈米粒子與配體的金硫鍵結可被任何帶有硫醇基的分子進行置換，並透過實驗確定配體的不穩定性主要源自於硫醇基分子的存在而非配體的自體氧化脫落。未來期望能將此膠體電泳檢測應用於奈米載體表面設計，作為細胞培養及動物實驗的前測試，使實驗進行奈米載體設計時有更好的選擇，能以簡單且快速的預測材料進入細胞或動物實驗後的穩定性趨勢，減少過度的資源浪費及無謂的犧牲，使生物醫學材料的檢測流程走向更精確快速且人道的未來。

關鍵字

膠體電泳 ； 金奈米粒子 ； 聚乙二醇化 ； 牛血清白蛋白 ； 半胱胺酸 ； 調理素化 ； 金硫鍵結 ； 快速感測 ； 環境老化測試

並列摘要

This study provides a new gel electrophoresis protocol for simple and rapid detection of nanoparticle stability which is always required for nanomedicine development. Our detection protocol can easily sense the density of Poly(ethylene glycol) ligands on AuNPs surface, ligand bonding stability and aging. We used cysteine as desorbents and BSA as opsonin to simulate the opsonization on PEGylated AuNPs, and used the gel electrophoresis to distinguish the stable and unstable AuNPs by inversed charge effect. The result showed that we successfully use BSA to detect the PEG density on the AuNPs surface, and combine the desorbents to force the opsonization of AuNPs, which clearly demonstrated the instability of PEGylated AuNPs. And the stability test reaction can be accelerated by heat treating. We have shown that electrophoretic detection can analyze the stability of AuNPs more effective than the UV-VIS spectrometer as well as the zeta potential analyzer. In addition, our heat treatment experiment has proved that ligands on AuNPs surface can be exchanged by any molecule that content thiol group. Different ligand designs do enhance the stability of gold nanoparticles that can slow down the opsonization of AuNPs. Finally, we give a general protocol that can easily detect the stability of AuNPs, and this method can be used on different sizes of PEGylated AuNPs. More importantly, the cost of this test is quite low but the results are relatively effective. We hope our study can give scientists a better choice that can easily evaluate the biological half-life of gold nanoparticle stability before cell culture and animal test, to avoid waste and unnecessary animal sacrifices.

並列關鍵字

electrophoresis ； gold nanoparticles ； Poly(ethylene glycol) ； PEGylated ； opsonization ； BSA ； cysteine ； rapid detection ； nanoparticle stability

參考文獻

參考文獻

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