隨著科學及醫療領域的持續發展，目前已有許多有別於傳統的檢測方法被提出並受到重視。其中，側向層析分析法（Lateral flow assay, LFA）是一種廣泛使用的即時檢測技術，在多個研究領域得到應用，例如：醫學診斷、環境監測、食品安全、農業獸醫應用及藥物檢測等。相較於其他現有的檢測方式，側向層析分析法具有簡易使用、快速檢測、低成本及具有可攜帶性等優點。然而此方法雖具有迅速且專一的特性，目前三明治型側向層析分析法的大多使用於大分子，例如抗原、蛋白質及核酸等，偵測小分子則較常使用競爭型側向層析分析法進行檢測，如此易導致結果判讀複雜性。於這篇論文中我們提出一種透過調控親和力的方式，並且基於側向層析分析平台的半合成蛋白開關，以增強訊號開啟方式來偵測有機小分子。此原理為透過目標小分子與探針上的配體競爭蛋白之結合位點，進而調控探針上之生物素與測試線上鏈黴親和素的結合能力，並透過特殊探針結構修飾使訊號明顯增強，達到提高靈敏度的效果。
在不含有目標小分子的樣品中，探針上生物素衍生物因周圍會產生較大的立體障礙，使生物素無法與測試線上之鏈黴親和素結合，呈現訊號關閉的狀態。當樣品中具有目標小分子，生物素周圍的空間立障會被消除，於流經測試線時會與鏈黴親和素結合，並因為探針上特殊結構修飾而使訊號放大，呈現訊號明顯開啟的狀態。基於這項研究設計，我們利用此訊號表達及增強機制、模塊化的探針特性及側向層析分析法的優點，成功實現偵測小分子的目的，並希望於未來擴大此技術的應用範疇以達到更多重要生物分子的即時檢測。

With the continuous development of science and medical treatment, many novel detection methods have been proposed and attracted much attention. Among them, Lateral flow assay (LFA) has been widely applied in many research fields, including medical diagnosis, environmental monitoring, food safety, agricultural and veterinary applications and drug detection. Compared with other detection methods, lateral flow assay has the advantages of simple use, rapid detection, low cost and portability. However, most of the sandwich-type LFA methods are used for macromolecules, such as antigens, proteins, and nucleic acids. Small molecules are more commonly detected by competitive type LFA, it is easy to lead to the result interpretation complexity. To detect small organic molecules by signal-on format, we propose an affinity-tunable semisynthetic protein switch based on lateral flow assay. The principle is regulating the binding affinity by utilizing the competition between the target small molecule and the ligands, and amplifying the signal through specific structural modification.
In the absence of target molecule, the biotin cannot combine with the streptavidin on the test line due to the large steric hindrance, so the signal is turned off. When the sample contains the target small molecule, the steric hindrance will be eliminated, and it will be captured by streptavidin when it flows through the test line. As a result, the signal will be turned on. Besides, the signal will be amplified due to the special structural modification so we can achieve lower detection limit. Based on this research design, we have successfully realized the purpose of detecting small molecules by taking advantage of semisynthetic protein switch and lateral flow assay, and believe that the application of this technology will be expanded to achieve more point-of-care detection of important biomolecules.

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