文献类型：期刊文献

英文题名：An AIE-based enzyme-activatable fluorescence indicator for Western blot assay: Quantitative expression of proteins with reproducible stable signal and wide linear range

作者：Zhou, Tijian;Wang, Qi[1,2];Liu, Ming;Liu, Zhenxing;Zhu, Zhirong;Zhao, Xiaolei;Zhu, Wei-Hong[1,2]

机构：[1]East China Univ Sci & Technol, Sch Chem & Mol Engn,Joint Int Res Lab Precis Chem, Frontiers Sci Ctr Materiobiol & Dynam Chem,Key La, Feringa Nobel Prize Scientist Joint Res Ctr,Shang, Shanghai 200237, Peoples R China;[2]East China Univ Sci & Technol, Feringa Nobel Prize Scientist Joint Res Ctr, Frontiers Sci Ctr Materiobiol & Dynam Chem, Sch Chem & Mol Engn,Inst Fine Chem,Joint Int Res, Shanghai 200237, Peoples R China

年份：2021

卷号：2

期号：2

外文期刊名：AGGREGATE

收录：EI(收录号：20233214485682);WOS:【ESCI(收录号:WOS:000710556400001)】；

基金：This work was supported by NSFC Science Center Program (21788102), NSFC Major Research Project (91959202), National Key Research and Development Program (2016YFA0200300), NSFC/China (21974047 and 21622602), Shanghai Pujiang Program (19PJ1402300), China Postdoctoral Science Foundation (2020M671328), Fundamental Research Funds for the Central Universities (222201814013), and Postdoctoral Science Foundation of Jiangsu Province (2020Z189).

语种：英文

外文关键词：aggregation-induced emission; amphiphilic fluorescence indicator; Western blot assay

摘要：Western blot is a commonly used experimental method to analyze the protein expression. However, the most commonly used chromogenic indicator based on chemiluminescence is limited by narrow linear range and unstable quantitative reproducibility, whereas the recently developed fluorescent indicator suffers from poor detection limit. Herein, we report an enzyme-activatable fluorescence indicator to quantify proteins with reproducible stable signal and wide linear range, through introducing the hydrophilic alkaline phosphatase (ALP)-triggered phosphoric acid moiety into our established aggregation-induced emission (AIE) building block of quinoline-malononitrile (QM). In this strategy, the indicator DQM-ALP disperses well in both aqueous and lipid environments to exhibit initial "off" fluorescence, but when exposing to the ALP-coupled secondary antibody on the PVDF membrane, the specific enzymatic turnover would liberate hydrophobic AIE luminogen (AIEgen) QM-OH to emit strong luminescence, thereby achieving an ideal "off-on" state for sensitively imaging proteins with high signal-to-noise (S/N) ratio. Moreover, benefiting from the excellent signal stability of AIE fluorophore, DQM-ALP indicator exhibits superior quantitative analysis of protein expression with high reproducibility. Upon taking advantage of the AIEgens to reduce high concentration-induced luminance quenching, the linear quantification range is extremely expanded. In contrast with the traditional chemiluminescent indicator, the AIE-based enzyme-activatable indicator DQM-ALP not only greatly improves the signal stability for quantitative reproducibility, but also expands the linear quantification range, and further provides a practical alternative reagent for fluorescence Western blot assay.