Zhang Xinfeng, Deng Li, Huang Chengpeng, Zhang Jinyi, Hou Xiandeng, Wu Peng, Liu Juewen
College of Material and Chemistry & Chemical Engineering, Chengdu University of Technology, Chengdu, 610059, P.R. China.
Analytical & Testing Centre, Sichuan University, Chengdu, 610064, P.R. China.
Chemistry. 2018 Feb 21;24(11):2602-2608. doi: 10.1002/chem.201704866. Epub 2018 Jan 9.
H O and horseradish peroxidase (HRP) are commonly used together in bioassays. HRP is required to accelerate the reaction between a chromogenic substrate (e.g., 3,3',5,5'-tetramethylbenzidine, TMB) and H O , and thus amplifies the signal. Herein, molecular oxygen for enzyme-free and H O -free oxidation is explored, still using the same colorimetric reaction. Restricted by spin selection rules, the ground-state triplet oxygen needs to be converted to the singlet state to oxidize TMB. Phloxine B (PB) is used as the photosensitizer because of its excellent performance and safety. Under green light irradiation, each PB has a turnover of approximately 51 TMB molecules in 20 min, making PB a "molecular enzyme mimic" for signal amplification. With its small size, multiple PB molecules are loaded on a graphene oxide nanosheet to design a modified enzyme-linked immunosorbance (ELISA) assay (termed photosensitization immunosorbent assay, PISA), improving the 1:1 enzyme/target ratio to n:1. PISA is more sensitive for carcinoembryonic antigen than a commercial ELISA kit, and successfully measures the antigen in the serum of multiple cancer patients. This simple and green method of oxidation coupled with the small size of the photosensitizer and graphene oxide may enable many other applications in biosensor development, smart materials, and energy harvesting.
过氧化氢（H₂O₂）和辣根过氧化物酶（HRP）常用于生物测定中。需要HRP来加速显色底物（如3,3',5,5'-四甲基联苯胺，TMB）与H₂O₂之间的反应，从而放大信号。在此，探索了用于无酶和无H₂O₂氧化的分子氧，仍使用相同的比色反应。受自旋选择规则限制，基态三线态氧需要转化为单线态以氧化TMB。由于其优异的性能和安全性，使用了孟加拉玫瑰红（PB）作为光敏剂。在绿光照射下，每个PB分子在20分钟内可使约51个TMB分子发生转化，使PB成为用于信号放大的“分子酶模拟物”。由于其尺寸小，多个PB分子负载在氧化石墨烯纳米片上，设计了一种改进的酶联免疫吸附测定法（称为光敏免疫吸附测定法，PISA），将1:1的酶/靶标比例提高到n:1。PISA对癌胚抗原的检测比商业ELISA试剂盒更灵敏，并成功检测了多名癌症患者血清中的抗原。这种简单且绿色的氧化方法，加上光敏剂和氧化石墨烯的小尺寸，可能会在生物传感器开发、智能材料和能量收集等许多其他应用中发挥作用。
