Activity of AC electrokinetically immobilized horseradish peroxidase.

Autor: Prüfer M; Fraunhofer Institute for Cell Therapy and Immunology, Branch Bioanalytics and Bioprocesses (IZI-BB), Potsdam-Golm, Germany., Wenger C; IHP GmbH - Leibniz Institute for Innovative Microelectronics, Frankfurt/Oder, Germany., Bier FF; Institute of Biochemistry and Biology, University of Potsdam, Potsdam-Golm, Germany., Laux EM; Fraunhofer Institute for Cell Therapy and Immunology, Branch Bioanalytics and Bioprocesses (IZI-BB), Potsdam-Golm, Germany., Hölzel R; Fraunhofer Institute for Cell Therapy and Immunology, Branch Bioanalytics and Bioprocesses (IZI-BB), Potsdam-Golm, Germany.
Jazyk: angličtina
Zdroj: Electrophoresis [Electrophoresis] 2022 Oct; Vol. 43 (18-19), pp. 1920-1933. Date of Electronic Publication: 2022 Aug 09.
DOI: 10.1002/elps.202200073
Abstrakt: Dielectrophoresis (DEP) is an AC electrokinetic effect mainly used to manipulate cells. Smaller particles, like virions, antibodies, enzymes, and even dye molecules can be immobilized by DEP as well. In principle, it was shown that enzymes are active after immobilization by DEP, but no quantification of the retained activity was reported so far. In this study, the activity of the enzyme horseradish peroxidase (HRP) is quantified after immobilization by DEP. For this, HRP is immobilized on regular arrays of titanium nitride ring electrodes of 500 nm diameter and 20 nm widths. The activity of HRP on the electrode chip is measured with a limit of detection of 60 fg HRP by observing the enzymatic turnover of Amplex Red and H 2 O 2 to fluorescent resorufin by fluorescence microscopy. The initial activity of the permanently immobilized HRP equals up to 45% of the activity that can be expected for an ideal monolayer of HRP molecules on all electrodes of the array. Localization of the immobilizate on the electrodes is accomplished by staining with the fluorescent product of the enzyme reaction. The high residual activity of enzymes after AC field induced immobilization shows the method's suitability for biosensing and research applications.
(© 2022 The Authors. Electrophoresis published by Wiley-VCH GmbH.)
Databáze: MEDLINE
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