小分子型蛋白質螢光探針近年來逐漸受到重視，其具有快速偵測、具專一性、高靈敏度、優化簡易、高信號雜訊比等優點，目前有許多小分子型螢光探針之發表，但其多為酶活性型探針，主要利用蛋白質本身之催化活性，進而達到其偵測目的，如醣苷酶(glycosidases)、蛋白酶(proteases)等酶活性蛋白質，但此手法若要偵測非酶活性蛋白質將受重重限制。
在此我們以環境敏感螢光分子與對蛋白質之親和性配體結合，並形成一對目標蛋白質具專一性偵測之螢光探針，利用蛋白質結合域之疏水性環境與外部環境之差異，使環境敏感螢光分子產生劇烈之螢光變化。在此，我們成功建立一套SBD型螢光探針模型，並成功應用於人類碳酸酐酶(hCAII)、胰蛋白酶(trypsin)及抗生物素蛋白(avidin)之專一性檢測，其螢光增益最高達17倍，並延伸應用於細胞膜上hCAII表達之成像及可調節親和性配體於數種磺胺藥物偵測。此外我們延伸SBD型探針之成功概念，以Cy5 rotor螢光分子作其概念延伸，雖其選擇性仍有待改良，但其嶄新之偵測模式及螢光分子本身之特性，仍值得我們深入研究及探討。

In recent years, small molecule fluorescent turn-on probes are getting attention as they allow for sensitive, simple, and specific detection with high signal-to-background ratios. Currently most of the small-molecule fluorescent turn-on probes are designed for monitoring enzyme activities, for example, glycosidases, proteases. Typically, their fluorescence turn-on mechanism is based on an enzymatic reaction with the chemical probes to convert a non-fluorescent substrate into a fluorescent product. On the other hand, the design of fluorescence probes for non-enzymatic proteins remains a challenging task.
Herein, we introduce a new type of fluorescent turn-on probes, where a small molecule ligand is conjugated to an environment-sensitive fluorophore, for the selective detection of both enzymes and non-enzymatic proteins. The fluorescent turn-on mechanism is based on the binding of the ligand to a hydrophobic ligand binding domain of the target protein whereby the close proximity to the hydrophobic environment can influence the environment-sensitive fluorophore to exhibit stronger fluorescence. We successfully established a SBD type model for specific detection of hCAII, trypsin and avidin with fluorescent turn-on ratios of up to 17-fold. We also demonstrated that these fluorescent probes can be employed to visualize carbonic anhydrase expressed on the cell surface, and applied on several sulfa drugs detection by tunable ligand affinity. Furthermore, we also extended our protein detection strategy, in Cy5 rotor model, the proteins selectivity remain to be improved, but the novel design and characteristics of our detection method is worth to be developed by adapting fluorescent molecular rotor based on Cy5.

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