Fluorometric and Colorimetric Dual-Readout Immunoassay Based on an Alkaline Phosphatase-Triggered Reaction

• Published in: Analytical chemistry (Washington) Vol. 91; no. 12; pp. 7828 - 7834
• Main Authors: Zhao, Jiahui, Wang, Shuang, Lu, Shasha, Liu, Guoyong, Sun, Jian, Yang, Xiurong
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 18.06.2019
• Subjects: Alkaline phosphatase; Calcium-binding protein; Chemistry; Colorimetry; Dephosphorylation; Dopamine; Enzyme-linked immunosorbent assay; Immunoassay; Mining; Myocardial infarction; Phosphatase; Resorcinol; Sodium; Sodium salts; Substrates; Troponin; Troponin I
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/acs.analchem.9b01553

Alkaline phosphatase (ALP) usually acts as a signal transmitter in enzyme-linked immunosorbent assay (ELISA); therefore, developing an attractive ALP activity assay, especially using a preferable substrate, would help improve the efficiency and convenience of ELISA in practical applications. Herein we have first prepared an original and creative substrate, named m-hydroxyphenyl phosphate sodium salt (m-HPP), with a desirable dephosphorylation site for ALP. On the basis of the ALP-catalyzed hydrolysis of m-HPP to resorcinol and its subsequent specific nucleophilic reaction with dopamine, we have exploited a fluorometric and colorimetric dual-readout ALP activity assay and ALP-based ELISA system. Under the employed experimental conditions, highly sensitive and specific assay of ALP and cardiac troponin I (cTnI) has been accomplished in a straightforward way. Furthermore, the commendable sensing performance of our proposed ELISA in the determination of the cTnI level in diluted human serum unambiguously illustrates great potential in the early diagnosis of acute myocardial infarction.