螢光共振能量轉移(Florescence Resonance Energy Transfer, FRET)技術可應用在分子檢測與探討分子間交互作用，但是FRET訊號通常很弱，需要放大訊號才能偵測到FRET訊號，本實驗以修飾有stretapvidin的量子點(Quantum dots, QD)作為螢光供體，捕捉接有螢光受體Alexa 647的單股DNA(ssDNA)進行FRET檢測。透過交流電荷動力學，如：交流電滲流(AC electro-osmosis, ACEO)、介電泳(Dielectrophoresis, DEP)及電場誘導偶極(FIDA)來集濃QD，再以QD上修飾的stretapvidin來捕捉修飾有biotin的ssDNA進行FRET分子檢測，試圖放大FRET訊號提升效率，增加FRET檢測的靈敏度。
本文第三章，首先我介紹了新的電極設計，依據學姐梁紫涵(2017)的實驗結果發現ACEO的Pumping作用，此作用有助於電極聚集捕捉奈米粒子，我改良設計四角電極來增加AC pumping的作用。在做FRET實驗前要先進行QD掃頻測試，選擇適當頻率進行FRET實驗，實驗結果發現在QD有形成串鏈的位置FRET效率可提高到60%，而沒有串鏈的位置效率則僅有40%，平均的FRET效率可達到30%。QD串鏈的形成是由於電場誘導偶極(Field Induced Dipole Attraction, FIDA)的結果，而FRET效率的提升我認為與QD形成一維的有序排列有關。
本文第四章，根據第三章的結果：QD有向序性的排列有助FRET訊號的提升。基於上述結果我選擇以長鏈雙股的λDNA作為骨幹，透過QD表面修飾的stretapvidin與λDNA上的biotin以間隔200個鹼基對鍵結，再以交流電場捕捉及拉伸λDNA使QD形成一維的FRET感測器。本章分三部分，第一部份我以不同頻率分別捕捉λDNA以了解λDNA的介電性質，第二部分進行修飾有QD之λDNA的掃頻，研究其受電場作用捕捉及拉伸的現象，第三部分根據掃頻結果進行FRET檢測。實驗結果發現FRET效率均在48%以上且最高可提高到80%，平均的FRET效率可達到60%，明顯比單只有QD時的30%更高，我猜測可能的原因是QD在一維線狀下Diffusion flux是以1/r的方式緩慢衰減；相較於球形Diffusion flux是以1/r2的方式衰減。QD-DNA在拉伸形成一維線狀的幾何形狀下，ssDNA較容易擴散到QD-DNA上鍵結率較高而有較高的FRET效率。這些實驗結果顯示定向組裝QD使QD有一維的有序排列，可以有效增強FRET效率，同時提升FRET在生物分子檢測中的實用性。
在第三章及第四章我已實現FRET效率的提升，我本文第五章我調降Target ssDNA的濃度來進行FRET靈敏度檢測，實驗結果發現，隨著Target ssDNA濃度的降低，FRET訊號、效率有下降的趨勢，ssDNA濃度可下探至1.67×10-12M。

A successful FRET detection would rely on whether weak FRET signals can be overcome by having FRET signals greatly amplified. In this thesis, I report a new strategy to promote FRET sensing. I employ stretapvidin-coated quantum dot (QD) as the FRET donor to probe target single stranded DNA (ssDNA) tagged with Alexa 647 as the FRET acceptor. In the experiment, we first concentrate QDs under AC electric fields with joint effects of AC electro-osmosis (ACEO), dielectrophoresis (DEP), and field induced dipole attraction (FIDA). The target ssDNAs can then be captured by QDs, capable of emitting intensified FRET signals with unusually high FRET efficiencies. The results reveal that the measured FRET efficiency is very sensitive to how QDs are aggregated. I speculate that the FRET enhancement could be a result of directional assembly of QDs due to FIDA.
I further extend the idea that directional assembly of QDs might help increasing FRET signals. I conjugate QDs along the backbone of long double stranded λDNA molecules. Having QD-DNAs trapped and stretched by AC fields, I can turn such QD-DNAs into 1-D FRET sensors, capable of amplifying FRET signals more efficiently. Using these QD-λDNA nanowires, I find that the FRET efficiency is at least 30% and can reach as high as 86%. The averaged FRET efficiency is 44%, higher than 30% for the pure QD case. I am still able to detect target ssDNAs reliably at the concentration as low as 1.67 picoM.

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