Dopamine as a polymerizable reagent for enzyme-linked immunosorbent assay using horseradish peroxidase

Sumed Yadoung; Shinichi Shimizu; Surat Hongsibsong; Koji Nakano; Ryoichi Ishimatsu

doi:10.1016/j.heliyon.2023.e21722

Dopamine as a polymerizable reagent for enzyme-linked immunosorbent assay using horseradish peroxidase

Heliyon. 2023 Oct 28;9(11):e21722. doi: 10.1016/j.heliyon.2023.e21722. eCollection 2023 Nov.

Authors

Sumed Yadoung¹, Shinichi Shimizu², Surat Hongsibsong^{1

3

4}, Koji Nakano², Ryoichi Ishimatsu⁵

Affiliations

¹ Environmental Science Program, Faculty of Science, Chiang Mai University, 50200, Thailand.
² Department of Applied Chemistry, Graduate School of Engineering, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka, 819-0395, Japan.
³ School of Health Sciences Research, Research Institute for Health Science, Chiang Mai University, Chiang Mai, 50200, Thailand.
⁴ Environmental, Occupational Health Sciences and Non-Communicable Diseases Center of Excellence, Research Institute for Health Sciences, Chiang Mai University, Chiang Mai, 50200, Thailand.
⁵ Department of Applied Physics, University of Fukui, 3-9-1 Bunkyo, Fukui, 910-8507, Japan.

Abstract

We demonstrate that dopamine can be used as a reagent for colorimetric enzyme-linked immunosorbent assay (ELISA) using horseradish peroxidase (HRP). Dopamine was able to be polymerized in the presence of HRP and H₂O₂, and black polydopamine was obtained after the enzymatic reaction. Because of the black color, the absorbance was significantly changed in the whole range of the visible light region. Here, an indirect competitive ELISA based on the polymerization of dopamine was performed to detect a fluoroquinolone antibiotic, enrofloxacin. The antibiotic is commonly used in livestock farming. The anti-antibiotics antibody was produced from egg yolk from chicken hens. In the visible range, sufficient absorbance changes of ∼0.4∼0.5 and a low background level for the ELISA response were obtained, and the 50 % inhibitory concentration value at 450 nm was determined to be 26 ppb. The performance of the indirect competitive ELISA based on the polymerization of dopamine was compared to that based on the oxidation of catechol because dopamine has a catechol skeleton. By the complex of HRP and H₂O₂, catechol can be oxidized to o-benzoquinone having a maximum absorption wavelength of 420 nm. It was shown that the absorbance change in the case of polydopamine was about 2.5 times higher than that of catechol, where the background levels were similar. This confirms that the polymerization of dopamine significantly enhanced the photosignal.

Keywords: Catechol; Catechol amine; Enzymatic reaction; Polymerization.