Wang Shumin, Huang Mengna, Hua Jianhao, Wei Lin, Lin Shen, Xiao Lehui
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research, Ministry of Education, Key Laboratory of Phytochemical R&D of Hunan Province, College of Chemistry and Chemical Engineering, Hunan Normal University, Changsha 410081, China.
State Key Laboratory of Medicinal Chemical Biology, Tianjin Key Laboratory of Biosensing and Molecular Recognition, College of Chemistry, Nankai University, Tianjin 300071, China.
Nanoscale. 2021 Mar 12;13(9):4946-4955. doi: 10.1039/d0nr09232k.
Alkaline phosphatase (ALP) as a necessary hydrolase in phosphate metabolism is closely related to various diseases. Ultrasensitive detection of ALP with a convenient and sensitive method is of fundamental importance. In this work, a fluorescence resonance energy transfer (FRET)-based single-particle enumeration (SPE) method is proposed for the quantitative analysis of ALP. This strategy is based on the effective fluorescence suppression by a polydopamine (PDA) shell on the surface of semiconducting polymer nanoparticles (SPNs). PDA with broadband absorption in the UV-vis region can serve as an excellent quencher for SPNs. However, ascorbic acid (AA), the product of the hydrolysis of 2-phosphate-l-ascorbic acid trisodium salt (AAP) in the presence of ALP, can effectively inhibit the self-polymerization of dopamine (DA) to form a PDA layer. Therefore, ALP can be accurately quantified by counting the concentration-related fluorescent particles in the fluorescence image. A linear range from 0.031 to 12.4 μU mL-1 and a limit-of-detection (LOD) of 0.01 μU mL-1 for ALP determination are achieved. The spiked recoveries for ALP determination in a human serum sample are between 90% and 108% with RSD less than 3.1%. In summary, this convenient and sensitive approach proposed here provides promising prospects for ALP detection in a complex biological matrix.
碱性磷酸酶（ALP）作为磷酸盐代谢中一种必需的水解酶，与多种疾病密切相关。采用便捷灵敏的方法对ALP进行超灵敏检测至关重要。在这项工作中，提出了一种基于荧光共振能量转移（FRET）的单粒子计数（SPE）方法用于ALP的定量分析。该策略基于聚多巴胺（PDA）壳对半导体聚合物纳米粒子（SPNs）表面荧光的有效抑制。在紫外-可见区域具有宽带吸收的PDA可作为SPNs的优良猝灭剂。然而，在ALP存在下，2-磷酸-L-抗坏血酸三钠盐（AAP）水解产物抗坏血酸（AA）可有效抑制多巴胺（DA）的自聚合以形成PDA层。因此，通过计数荧光图像中与浓度相关的荧光粒子可准确量化ALP。实现了ALP测定的线性范围为0.031至12.4 μU mL-1，检测限（LOD）为0.01 μU mL-1。在人血清样品中测定ALP的加标回收率在90%至108%之间，相对标准偏差小于3.1%。总之，本文提出的这种便捷灵敏的方法为在复杂生物基质中检测ALP提供了广阔前景。
