應用大氣水霧電漿輔助快速固定蛋白質於生物感測器製程

Abstract

近年來，大氣常壓電漿沉積聚合技術已相當成熟，比起一般的低壓電漿，大氣常壓電漿不僅擁有較多的發展空間且花費的成本較低。本研究的目標為使用大氣常壓電漿沉積聚合將蛋白質固定至表面，並運用於生物感測器的製程。傳統固定蛋白質的方法不僅花費相當多的時間與試劑，而且步驟相當繁瑣，本研究所使用的新技術為大氣水霧輔助電漿沉積，不僅在表面產生羥基官能基，同時將蛋白質嵌入至薄膜，相較於傳統固定蛋白質方法更為快速，而且能避免使用有毒試劑。 大氣水霧輔助電漿沉積系統由兩平板陶瓷介電質所組成，介電質板上下兩端貼有鋁膠帶作為電極。實驗中所使用之蛋白質為牛血清白蛋白(Bovine serum albumin, BSA)，設定之工作電壓及頻率分別為3 kV與4 kHz，作為工作氣體氦氣之流量為5 SLM，白蛋白水溶液藉由通過霧化器形成水霧狀，進入至腔體內部前與乙烯混合。電漿聚合過程中，乙烯透過電漿解離並重新聚合成聚乙烯薄膜沉積於基材表面，水分子將解離成羥基官能基與聚乙烯薄膜鍵結，最終BSA將嵌入至聚乙烯薄膜內完成沉積聚合。 實驗結果分析，透過傅里葉轉換紅外光譜(FTIR)，波峰的疊加可以觀察出BSA成功地被固定到聚乙烯薄膜於矽晶圓上，以及被沖洗後薄膜殘留的訊號；藉由掃描式電子顯微鏡(SEM)觀察表面薄膜與BSA數量分佈的情況；最後藉由酵素結合免疫吸附分析法(ELISA)以及螢光蛋白質的染色，對被固定的BSA進行活性與螢光強度分析。藉由以上結果證明利用水霧輔助大氣電漿沉積BSA/聚乙烯薄膜，於薄膜附著性與蛋白質活性上均有良好的結果，可應用於生物感測器之製程。

Atmospheric pressure plasma deposition(APPD) polymerization has more possible applications and is less expensive when compared with low-pressure plasma polymerization. Protein immobilization is one of the applications used in biosensor fabrication. Compare with plasma immobilization, the standard process is not only time and reagent consuming, but also complicated. A novel technique, aerosol-assisted dielectric barrier discharge atmospheric pressure plasma deposition (DBD-APPD), can deposit hydroxyl functional groups and embed protein simultaneously, which makes the process faster and avoids using harmful reagent. The aerosol-assisted DBD-APPD system was constructed with two parallel ceramic plates and aluminum tape pasted on the center as electrode. The bovine serum albumin (BSA) was used in this experiment. The consuming power voltage was sinusoidal wave at 3 kV and frequency at 4 kHz. Total carrier gas helium was fixed at 5 SLM. The BSA solution was turned into aerosol while guiding helium flowing through an atomizer, then mixed with ethylene before entered the chamber. In the process of plasma polymerization, ethylene was disassociated by plasma then polymerized into PE film and deposited onto substrate. Simultaneously, the OH functional groups were disassociated from water and formed on PE film. And the protein molecules embedded in PE film. The surfaces properties of substrate were characterized by FTIR. We could found that the BSA signal appeared when we fed the BSA into the system. The SEM images showed the immobilized BSA on substrate. The protein activity was analyzed with ELISA assay. As a result, we immobilized BSA with aerosol-assisted DBD-APPD, and the result shows that the biosening application is feasible.