| 摘要: | 生物感測器在目前醫學領域尤其是轉譯醫學中扮演著十分重要的角色，生物感測技術為基礎醫療研究以及臨床診斷治療提供了一種快速簡便的檢測方法，並且其具有專一、靈敏、反應時間短等特性，運用在檢測微量生物指標(Biomarker)，期望能夠達到快速的疾病診斷之感測、少量檢體檢驗感測及高靈敏生醫感測等目標，以便提供醫療人員能夠快速方便使用的檢測平台，進而提升檢測效率，創造病人福祉。
自人類基因體計畫(Human Genome Project)啟動以來，基因資訊的快速獲得極充分利用，將人們朝向精確醫學(Precision Medicine)所強調之個人化預防醫學與治療邁進，核酸類似物的發展促使基因能更準確極方便的定序，從生命體最根源的基因表現來達到疾病的預防及治療，更進一步地將每個人的基因進行定序，規劃屬於個人專屬的健康照顧，將醫療推向另一個新的層級。
近年發展的核酸適體(aptamer)做為生物感測，其優點為方便取得，且能夠方便的改質上不同的官能基，使其應用面更為廣闊。
在本研究中運用的核酸適體(aptamer)為RNA-18核酸適體，可發展出一種具有辨識能力且放大訊號的檢測平台。
實驗分為三個階段，第一階段為核酸適體之合成；而後經由粒徑分析儀(DLS)來判定合成之大小。
第二階段為核酸適體的專一性結合測試，判定aptamer的結合能力是否正常。
第三階段在檢測平台上利用aptamer，運用於實際檢測物之檢測。
本實驗中藉由檢測平台來證實此核酸適體可成為新一代通用之訊號放大器，其靈敏性及專一性極佳，擁有便宜、合成程序少、穩定度高等優點，可預測是否罹患病症，發展快速簡便的檢測方法，在未來醫學領域必能發展一項便利又具可行性的檢測方法。
另一部分是本實驗室建立不帶電核酸於寡核苷酸(nDNA)合成改質之原則。此原則之建立以協助本實驗室於中性核酸探針改質以調整與目標核酸雜交之溶解溫度(Tm)與熱穩定性為主。利用不同的量測平台，能夠幫助我們了解nDNA與目標分子的Tm值，並且歸納出相關的設計原則，以利往後能夠發展高專一性、高靈敏性的探針能夠使用於檢測平台上，得到高效率的檢測。
;This research is separate into two part of experiments, First part is R18 RNA aptamer for a universal signal amplifying platform on ELISA, and second part is design nDNA in the sequence for improve melting temperature and have higher specificity, selectivity, and heat stability.
Enzyme-linked immunosorbent assay (ELISA) system is a useful detection technique for antigen or other biomolecules by using antibodies, but there also have some difficulties. For example, antibody is time-consuming in produce, expensive, difficult to label, and more background signal by labeling on secondary antibody, etc. However, development with the SELEX method which can select the nucleic acid based probe-aptamer. This molecule has more advantages than antibody. Aptamer is an artificial product which has high affinity and high specificity with target molecular. We find that R-18 RNA aptamer can become a signal amplifying platform. Finally, we use RNA aptamer to create a universal signal amplifying platform.
At the second part, we develope a neutralized DNA called “nDNA”, it backbone has no charge because of a methyl modified at the phosphate side. It can increase the specificity and selectivity with the target sequence, because of no charge backbone can help it to hybridize with target sequence in low salt solution. We design the position of the nDNA in the sequence and use many instrument to measure the melting temperature, afterthat we find nDNA can increase the melting temperature and heat stability with different position and numbers. Finally, we have some rules for how to design nDNA in the sequence, for high specificity and heat stability. We can use these rules to control melting temperature on the same platform afterthat we can use nDNA for more efficiency in diagnosis or therapeutic applications. |
