整合磁性鎳奈米粒子與微流體晶片架設微流道酵素免疫檢測平台 - 以肺炎鏈球菌檢測為例

Abstract

酵素免疫分析法 ( Enzyme-linked Immuno-sorbent Assay, ELISA ) 是利用抗原與抗體間的專一性結合，搭配酵素呈色反應對樣本檢體進行檢測，此方法廣泛的應用在生物檢測領域。但因此技術特性需要繁複的液體操作與較長的反應時間，故仍有改善空間。而本研究整合磁性鎳奈米粒子與微流體晶片，架設微流道酵素免疫檢測平台，提供一個簡化操作的方式。在此以肺炎鏈球菌檢測為例，首先以化學還原法製備出具有順磁性的鎳/氧化鎳磁性奈米粒子作為載體，以帶有組胺酸標籤 ( his-tagged ) 的 Gβ1 蛋白，結合對肺炎鏈球菌表面之特定蛋白質 ( SMU290 ) 有辨認性的抗體，抓取肺炎鏈球菌，並利用微流體晶片進行檢測。此晶片具有三層夾層，基底的玻璃透過表面修飾上親、疏水間層，結合中層 PDMS與上層 PMMA 夾擠固定不同溶液，搭配用Arduino 控制板與馬達驅動 IC 的自製平台，以永久性強力磁鐵帶動磁性鎳奈米粒子達到微流道酵素免疫分析法的檢測。此酵素免疫檢測平台能降低操作時間與人工移液的誤差，輕便具可攜帶性、低成本耗材又可以自動化的檢測肺炎鏈球菌，其應用更可以延伸到任何與酵素免疫分析法有關的系統，以供偏遠區域及未來臨床應用。

The enzyme-linked immune-sorbent assay (ELISA) is a test that uses antibodies recognition and enzymes catalytic activity to identify a substance, and has been widely used as a diagnostic tool in clinical. However, performing an ELISA require various liquid handling steps and long binding time. In order to solve this problem, we have developed a micro fluidic platform, involving the application of magnetic nanoparticles, which can streamline the process of an ELISA in a fully automated manner for Streptococcus pneumoniae detection. First, we synthesized the paramagnetic surface-oxidized nickel nanoparticles (Ni/NiO NPs) to carry the His-tagged protein Gβ1. Then we assembled the SUM290 specific antibody on protein Gβ1. Lastly we integrated the nanoparticles on the micro fluidics chip for S. pneumoniae detection. The chip contains three different layers to trap the solutions; the bottom layer SiO2 has patterned on hydrophobic polymer, integrated with the middle layer PDMS and top layer PMMA. With Arduino chip and motor IC, we developed an automated platform for S. pneumoniae detection. Micro fluidics ELISA can reduce the manual handling and operation time. Furthermore, the develop system can be extended to multiple areas for ELISA-related assays. This economic, rapid and portable system may become a promising platform for sensing S. pneumoniae in the clinical application.