Guangzhou Key Laboratory of Analytical Chemistry for Biomedicine, School of Chemistry, South China Normal University, Guangzhou, Guangdong 510006, People's Republic of China.
State Key Laboratory for the Chemistry and Molecular Engineering of Medicinal Resources, College of Chemistry and Pharmaceutical Science, Guangxi Normal University, Guilin, Guangxi 541004, People's Republic of China.
Anal Chem. 2022 May 10;94(18):6711-6718. doi: 10.1021/acs.analchem.1c05463. Epub 2022 Apr 29.
Alkaline phosphatase (ALP), an essential hydrolase with crucial roles in living organisms, has widely been regarded as a biomarker for various human diseases in clinical diagnoses. Herein, taking advantage of cobalt oxyhydroxide (CoOOH) nanoflakes and nonenzymatic cascade recycling amplification (CRA), a highly sensitive and label-free fluorescence biosensing strategy for the determination of ALP activity is introduced. In our design, ALP can promote the dephosphorylation of l-ascorbic acid 2-phosphate (AAP) to reduce ascorbic acid (AA), which is then able to decompose CoOOH in a nucleic acids@CoOOH nanocomplex into Co cofactors. Further, enzyme-free CRA was rapidly initiated by integrating DNAzyme recycling amplification and catalytic hairpin assembly, resulting in the generation of an abundance of G-quadruplex structure-contained DNA duplexes. In the presence of thioflavin T (ThT), analytical target ALP was converted in an amplified and activatable fluorescence signal. The experimental results show that this method can be applied for the quantitative analysis of ALP activity with a low detection limit of 0.027 mU/mL. Moreover, this developed biosensing approach exhibits excellent specificity, and the evaluation of ALP activity in the complex human serum samples was successfully realized, indicating that it can afford a reliable, robust, and cost-effective nanoplatform for an ALP-based clinical diagnosis and for biomedical research.
碱性磷酸酶（ALP）是一种重要的水解酶，在生物体内具有关键作用，已广泛被认为是临床诊断中各种人类疾病的生物标志物。在此，我们利用钴氢氧化物（CoOOH）纳米片和非酶级联循环扩增（CRA），提出了一种用于测定 ALP 活性的高灵敏、无标记荧光生物传感策略。在我们的设计中，ALP 可以促进 l-抗坏血酸 2-磷酸（AAP）的去磷酸化，从而还原抗坏血酸（AA），AA 随后能够将核酸@CoOOH 纳米复合物中的 CoOOH 分解为 Co 辅因子。此外，通过整合 DNA 酶循环扩增和催化发夹组装，快速引发无酶 CRA，从而产生大量包含 G-四链体结构的 DNA 双链。在硫黄素 T（ThT）存在下，分析物 ALP 被转化为放大和激活的荧光信号。实验结果表明，该方法可用于定量分析 ALP 活性，检测限低至 0.027 mU/mL。此外，这种开发的生物传感方法表现出优异的特异性，并成功实现了复杂人血清样品中 ALP 活性的评估，表明它可以为基于 ALP 的临床诊断和生物医学研究提供可靠、稳健且具有成本效益的纳米平台。
