本研究設計出一連續式自動化微流體晶片系統，藉由系統性配位子指數增益演繹程序(SELEX)，針對甲型胎兒球蛋白(alpha-fetoprotein)，進行以磁珠為固相的適合體篩選流程。將磁珠應用於適合體之萃取，並在晶片上進行聚合酶連鎖式反應增幅，可以減少人為操作的誤差，更大幅縮短操作時間與步驟，具有低樣品及檢體消耗量、低耗能、體積小以及成本低等優點，相較傳統技術，有著突破性的發展價值。
本微流體晶片系統中包含兩種微型幫浦、微混合器以及微型聚合酶連鎖反應模組等。其中線型微型幫浦在通入氣壓為10 psi，操作條件為10 Hz 時，能夠準確地傳輸檢體 (流速為1 μL/sec)，可避免檢體或試劑的浪費；而環狀微型幫浦在同樣氣壓下，操作條件為15 Hz 時，兼具混合與大量傳輸的功能 (流速為21.7 μL/sec)；微混合器可於5秒內達到約98.6 % 的混合效果，可使檢體於最短的時間內有最佳的混合效率；微型聚合酶連鎖反應模組之試劑儲存區的溫度低於40℃，可確保酵素與蛋白質的活性不被破壞，並能成功執行 DNA 之增幅。而所有元件將整合於一微型晶片系統上，可達到自動化傳輸流體、混合的功能，並於最後將微流體系統晶片應用於甲型胎兒球蛋白適合體篩選流程。
本研究成功地達到連續式的系統性配位子指數增益演繹程序篩選流程，單一次流程只需花費約六十分鐘，整套篩選流程(五次)花費時間只需近六小時即可完成。同時使用此套系統進行甲型胎兒球蛋白適合體篩選，實驗結果也驗證相關的適合體可被萃取出來，而本適合體的鑑結強度藉由與另一已知解離常數 (KD=2.37 nM) 適合體來進行比較評估，顯示其具有相當的親和力，來進行甲型胎兒球蛋白的檢測。

This study reports a continuous microfluidic system which utilizes systematic evolution of ligands by exponential enrichment (SELEX) technique for rapid screening of specific aptamers for alpha-fetoprotein (AFP). The system utilizing magnetic bead-based techniques to select DNA aptamers has several advantages over its large-scale counterparts, including rapid screening process, automation, and less consumption of samples/reagents. With a microfluidic control module for incubation process and an on-chip polymerase chain reaction (PCR) module for amplification of these selected aptamers, the entire SELEX process can be automatically performed in a shorter period of time.
This new microfluidic system integrates two types of micropumps with normally-closed microvalves, a circular-shape micromixer and an on-chip PCR module on a single chip. The linear-shaped micropump has been adopted to achieve a low flow rate (1 μL/sec) to control the PCR reagent transportation accurately. The circular-shape micropump with a high flow rate (21.7 μL/sec) was used for rapid washing process. The circular-shape micromixer can incubate the DNA with magnetic beads efficiently. The mixing index was found to increase from 16.13 % to 98.6 % within five seconds. The PCR module can rapidly amplify the target DNA. Furthermore, the temperatures of the PCR reagents and magnetic beads reservoirs was kept at 40℃ or lower during the PCR procedure.
In this study, the continuous operation of the SELEX process has been achieved in screening specific aptamers of AFP. Compared to the traditional SELEX process, this continuous microfluidic system is efficient and consumes fewer sample volumes. It took approximately 5 hours for a SELEX process with five screening runs, which was faster than that of a traditional SELEX process (8 hours for 5 rounds). And the binding affinity of this specific aptamer was compared with another aptamer which had been measured by a BIAcore X system with the dissociation constant (KD) 2.37 nM. The results show that the specific aptamer may has affinity for further AFP detection. This continuous microfluidic system may provide a powerful platform for rapid screening of target- specific aptamers.

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