利用滾動循環擴增技術之細胞激素陣列化螢光分析訊號放大與最適化探討

Fluorescent-Signal Amplification and Optimization of Cytokine Array Using the Rolling-Circle Amplification Technique

葉庭秀(Ting-Shiu Ye)

指導教授 ： 吳瑞璋

中原大學/工學院/化學工程學系/碩士(2007年)

https://doi.org/10.6840/cycu200700521

摘要

中文摘要
本研究之蛋白質晶片是以偵測引起細胞增殖、變異、發炎等一系列免疫反應之微量細胞激素晶片。經環氧基改質之1”e3”玻片，在鋪設一級抗體後，以三明治免疫反應(immunoassay)捕捉抗原-人類白血球間質素-6 (Interleukin 6)，再利用以生物素(biotin)修飾之報告抗體-老鼠抗人類白血球間質素-6 (biotinylated mouse anti human IL6)，以鏈菌素(streptavidin)做為一架橋中間物來連結上另ㄧ修飾有生物素之環狀DNA片段，以滾動循環DNA擴增技術(Rolling-circle Amplification)複製出大片段之單股DNA產物，再雜合(hybridization)帶螢光修飾之單股DNA，最後經掃描螢光訊號得到免疫測試法之偵測結果。此訊號放大技術可用來降低晶片捕捉目標蛋白質之濃度，尤其針對微量即可造成嚴重反應的低濃度細胞激素-白血球間質素-6 (IL-6)，其蛋白質晶片(cytokine protein chip)之製備即為此研究之主要目的。本研究詳細敘述滾動循環擴增技術製備環狀模板之程序及測試方法，並指出細胞激素晶片製備時所面對之挑戰。

關鍵字

蛋白質晶片 ； 滾動循環

並列摘要

Abstract
This research developed a protein biochip to detect trace human cytokines, which are important to small-secreted proteins mediating and regulating immunity, cellular differentiation and proliferation, and inflammation in response to an immune stimulus. A sandwich immunoassay was built on 1”e3” glass slides, of which surface was modified with epoxy and a primary antibody to capture the target, human cytokine IL-6. A circular DNA template was attached onto the crystal-consistent domain of the report antibody. Composed with the Rolling Circle Amplification (RCA) technique, this cytokine biochip was developed and read out by fluorescent labels. This study states the preparation process and the characterization of the circular-DNA template in detail and also points out the challenges during the development.

並列關鍵字

rolling circle amplification ； protein microarray

參考文獻

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