可控化矽烷化:以巰基氮矽三環之硫醇-烯加成反應發展可功能化之抗沾黏生物介面

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姓名

粘佩儒(Pei-Ju Nien) 查詢紙本館藏

畢業系所

生醫科學與工程學系

論文名稱

可控化矽烷化:以巰基氮矽三環之硫醇-烯加成反應發展可功能化之抗沾黏生物介面
(Controlled Silanization: Functional Antifouling Biointerfaces Developed via Thiol-ene Chemistry with Mercaptosilatrane)

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摘要(中)

由於生物感測器的作業環境複雜，非特異性吸附現象往往影響生物感測器訊號判讀，因此本研究利用表面工程技術降低生物感測器介面的非特異性吸附，同時提供可功能化修飾的特性，化學接枝生物識別分子(bio-recognition elements)，以提高生物感測器的訊噪比(signal-to-noise ratio, S/N ratio)。矽烷官能基(silane)常用於形成自組裝薄膜(self-assembled monolayer, SAM)，然而因其容易水解，常會導致分子團聚和表面修飾不均勻。相反地，氮矽三環結構(silatrane)因存在氮和矽之間的分子內相互作用，而具有較高的化學穩定性，因此可以控制矽烷化過程並提高均勻性，藉此提升感測器的靈敏度與再現性。在這項研究中，2-甲基丙烯醯氧基乙基磷酸膽鹼(2-methacryloyloxyethyl phosphorylcholine , MPC)和帶有羧酸基的聚乙二醇(poly(ethylene glycol), PEG)將通過硫醇-烯加成反應(thiol-ene reaction)與巰基氮矽三環(mercaptosilatrane, MPS)聚合。MPC和PEG具有出色的生物相容性和抗非特異性吸附能力，此外，PEG上的羧基可以轉化為可功能化的中間體以產生具有生物選擇性的表面。在抗沾黏測試中，表面塗佈塗層MPS-MPC5後，細菌與蛋白質的貼附量明顯地降低。在矽烷官能基與氮矽三環的表面型態比較中，氮矽三環的表面呈現較佳的均勻性。接著，使用MPS-MPC3-carboxylated PEG2進行生物感測器的測試。光纖式粒子電漿共振感測儀(fiber optic particle plasmon resonance, FOPPR)是利用奈米金粒子獨特光學性質與光纖全反射特性組成的高靈敏生物檢測系統。將MPS-MPC3-carboxylated PEG2修飾於感測區後可降低其非特異性吸附現象，並且在多濃度測試中，從2.5 × 10-13 g / mL至2.5 × 10-8 g / mL有可靠的線性響應，其偵測極限為3.1 × 10-14 g / mL。此新型的共聚物為基材表面帶來較佳的均勻性、良好的抗汙效果及可功能化的特性，具有極大的潛力，能因應市場需求、促進生物感測器的發展。

摘要(英)

Surface modification for biosensors is of importance for improving specificity and sensitivity, particularly for the detection in complex medium. Modifier, silane, is commonly used to form a self-assemble monolayer (SAM). However, silane molecules are very sensitive to moisture and environmental conditions, resulting in agglomeration and rough surface. Very recently, we demonstrated that silatrane is chemically stable to hydrolysis because of its transannular bond between nitrogen and silicon. Therefore, the silanization procedure is controlled and the homogeneity is improved by using silatranes. In this study, 2-methacryloyloxyethyl phosphorylcholine (MPC) and carboxylated poly(ethylene glycol) (carboxylated PEG) will be chemically attached to mercaptosilatrane (MPS) via thiol-ene reaction. MPC and carboxylated PEG provide excellent biocompatibility and high resistance to nonspecific adsorption. Besides, the carboxyl group can be transformed into functional intermediate for conjugation of bio-recognition elements to develop a sensing surface. In an antifouling test, both of bacterial adhesion and protein adsorption on the surface were reduced significantly after modification of MPS-MPC5. In the comparison of silane and silatrane, silatrane showed better homogeneity than silane. Next, MPS-MPC3-carboxylated PEG2 was synthesized to perform molecular detection. Fiber optic particle plasmon resonance (FOPPR) is a rapid and ultrasensitive biosensing device based on fiber optic nanogold-linked immunosorbent assay. It showed the lower nonspecific adsorption after MPS-MPC3-carboxylated PEG2 modified the detection region. For the multiple concentration test, it provides a wide linear response range from 2.5×10-13 g/mL to 2.5×10-8 g/mL (6 orders) and an extremely low limit of detection of 3.1×10-14 g/mL for NT-proBNP(1-76), which is an important biomarker for heart failure. The new type of functional silatrane assemblies offers antifouling properties, great homogeneity, and functionalized capacity. It has great potential to improve the development of biosensors.

關鍵字(中)

★ 可控矽烷化
★ 非特異性吸附
★ 生物感測器介面
★ 雙離子材料
★ 聚乙二醇

關鍵字(英)

★ controllable silanization
★ nonspecific adsorption
★ biosensor interface
★ zwitterionic materials
★ poly(ethylene glycol)

論文目次

中文摘要 i
Abstract ii
致謝 iv
目錄 v
圖目錄 viii
表目錄 xi
一、文獻回顧 1
1-1　　生物感測器 1
1-1-1　影響生物感測器訊號之因素 2
1-2　　自組裝抗汙塗層 3
1-2-1　矽烷官能基 4
1-2-2　氮矽三環類化合物 9
1-3　　抗非特異性吸附之材料 14
1-3-1　非特異性吸附現象 14
1-3-2　抗沾黏材料特性 15
1-3-3　聚乙二醇材料 15
1-3-4　雙離子材料 15
1-3-5　PC類雙離子材料 16
1-4　　硫醇-烯加成反應 18
1-5　　生物辨識元件之固定化流程 20
1-6　　光纖式奈米生物感測儀 22
二、研究目的 23
三、實驗藥品與設備及實驗方法 24
3-1　　實驗藥品與設備 24
3-2　　材料合成 27
3-2-1　巰基氮矽三環(MPS) 27
3-2-2　聚乙二醇羧酸化之過程 27
3-2-3　MPS-MPCn-carboxylated PEGm 28
3-2-4　無水甲醇之製備 29
3-3　　實驗方法 29
3-3-1　自組裝分子膜之製備 29
3-3-2　衰減全反射傅立葉轉換紅外線光譜(Attenuated total reflectance-fourier transform infrared spectra，ATR-FTIR)鑑定 29
3-3-3　水接觸角之量測 30
3-3-4　薄膜厚度之量測 30
3-3-5　表面型態之測量 30
3-3-6　表面元素鑑定 30
3-3-7　細菌貼附測試 31
3-3-8　蛋白質貼附測試 31
3-3-9　以傳統三明治法偵測目標物 32
3-3-10　光纖式粒子電漿共振奈米生物感測器(Fiber optic particle plasmon resonance，FOPPR)偵測目標物 32
四、結果與討論 34
4-1　　以MPS-MPCn比較分子鏈長對修飾效果之影響 34
4-1-1　MPS-MPCn之NMR光譜圖 34
4-1-2　MPS-MPCn之薄膜接觸角量測 36
4-1-3　MPS-MPCn之薄膜厚度量測 36
4-1-4　MPS-MPCn之抗細菌沾黏測試 37
4-1-5　MPS-MPCn之抗蛋白質沾黏測試 39
4-2　　比較MPS-MPC5與MPTMS-MPC5 41
4-2-1　MPTMS-MPC5之NMR頻譜分析 41
4-2-2　MPS-MPC5與MPTMS-MPC5之水接觸角比較 42
4-2-3　MPS-MPC5與MPTMS-MPC5之抗生物分子沾黏測試 42
4-2-4　MPS-MPC5與MPTMS-MPC5之表面型態分析 44
4-2-5　MPS-MPC5與MPTMS-MPC5之薄膜厚度比較 45
4-3　　以MPS-MPCn-carboxylated PEGm偵測目標物 46
4-3-1　Carboxylated PEG之NMR圖譜分析 46
4-3-2　Carboxylated PEG之FTIR圖譜分析 47
4-3-3　MPS-MPCn-carboxylated PEGm表面元素分析 48
4-3-4　探討表面的活性位(active site)比例對目標物訊號影響 50
4-3-5　探討目標物濃度對偵測訊號之影響(ELISA) 51
4-3-6　以FOPPR測試抗非特異性吸附能力 52
4-3-7　以FOPPR進行多濃度測試 53
五、結論 56
六、未來展望 57
七、參考文獻 58

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指導教授

黃俊仁(Chun-Jen Huang)

審核日期

2020-1-18

推文