Developing a fluorescence substrate for HRP-based diagnostic assays with superiorities over the commercial ADHP  

作　　者：Zhichao Zhou Fuqian Chen Xiaotong Xia Dong Ye Rong Zhou Lei Li Tao Deng Zhenhua Ding Fang Liu 

机构地区：[1]Department of Radiation Medicine,Guangdong Provincial Key Laboratory of Tropical Disease Research,School of Public Health,Southern Medical University,Guangzhou 510515,China [2]School of Pharmaceutical Sciences,Guangzhou University of Chinese Medicine,Guangzhou 510006,China [3]Artemisinin Research Center,Guangzhou University of Chinese Medicine,Guangzhou 510006,China [4]State Key Laboratory of Respiratory Disease,the First Affiliated Hospital of Guangzhou Medical University,Guangzhou Medical University,Guangzhou 510120,China [5]Guangzhou Laboratory,Bio-Island,Guangzhou 510030,China [6]Nanshan Pharmaceutical Innovation Research Institute of Guangdong Province,Foshan 528225,China

出　　处：《Chinese Chemical Letters》2024年第6期439-444,共6页中国化学快报（英文版）

基　　金：the funding support from Key-Area Research and Development Program of Guangdong Province(No.2022B1111020003);Guangzhou Talents Program for Innovation and Entrepreneurship(No.2021-L010);the Foshan“Blue Ocean Talent Program”for Innovation and Entrepreneurship(No.2230032002063)。

摘　　要：The combination of horseradish peroxidase(HRP)and a fluorescence substrate has been attracting great interests in developing sensitive biochemical analysis and immunoassays.10-Acetyl-3,7-dihydroxyphenoxazine(ADHP or Amplex red)is the most sensitive fluorogenic substrate known for HRP in current market,however,it suffers from some drawbacks,such as non-specific reactivity to carboxylesterase and limited fluorescence stability.In the present study,a novel HRP substrate10-cyclopropylcarbonyl-dichloro-dihydroxyphenoxazine(AR-2),has been prepared,which exhibited improved sensitivity than ADHP in sensing HRP.Moreover,the fluorescence of AR-2/HRP demonstrated improved tolerance to physiological relevant p H fluctuation as compared to ADHP/HRP.Successful detection of uric acid/urate oxidase reaction indicated excellent application prospect of AR-2/HRP for monitoring H_(2)O_(2)-generating biochemical reactions.More interestingly,an enzyme-linked immunosorbent assay(ELISA)using AR-2 as the fluorescence reporter has been successfully used in detecting IgG against severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)from human serum samples.Overall,AR-2 exhibits improved performances over the commercial ADHP,which will be an ideal alternative to ADHP in HRP-based fluorescence biochemical analysis and immunoassays.

关 键 词：Fluorescence immunoassay HRP fluorescence substrate Hydroresorufin Amplex red/ADHP SARS-CoV-2 

分 类 号：Q503[生物学—生物化学] O657.3[理学—分析化学]