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Activity of AC electrokinetically immobilized horseradish peroxidase

  • Dielectrophoresis(DEP) is an AC electrokinetic effect mainly used to manipulate cells.Smaller particles,like virions,antibodies,enzymes,andevendyemolecules can be immobilized by DEP as well. In principle, it was shown that enzymesare active after immobilization by DEP, but no quantification of the retainedactivity was reported so far. In this study, the activity of the enzyme horseradishperoxidase (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 H2O2 to fluorescent resoruf in 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 electrodesis accomplished by staining with theDielectrophoresis(DEP) is an AC electrokinetic effect mainly used to manipulate cells.Smaller particles,like virions,antibodies,enzymes,andevendyemolecules can be immobilized by DEP as well. In principle, it was shown that enzymesare active after immobilization by DEP, but no quantification of the retainedactivity was reported so far. In this study, the activity of the enzyme horseradishperoxidase (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 H2O2 to fluorescent resoruf in 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 electrodesis accomplished by staining with the fluorescent product of the enzyme reac-tion.The high residual activity of enzymes after AC field induced immobilization shows the method’s suitability for biosensing and research applications.show moreshow less

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Metadaten
Author: Mareike Prüfer, Christian WengerORCiD, Frank F. Bier, Eva-Maria Laux, Ralph Hölzel
DOI:https://doi.org/10.1002/elps.202200073
ISSN:1522-2683
Title of the source (English):Electrophoresis
Document Type:Scientific journal article peer-reviewed
Language:English
Year of publication:2022
Tag:dielectrophoresis; immobilization; nanoelectrodes
Volume/Year:43
Issue number:18-19
First Page:1920
Last Page:1933
Faculty/Chair:Fakultät 1 MINT - Mathematik, Informatik, Physik, Elektro- und Informationstechnik / FG Halbleitermaterialien
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