Assessment of Deoxyribonuclease Activity Using DNA Molecules Immobilized Between Microelectrodes
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https://doi.org/10.1007/s11664-020-08596-8 Ó 2020 The Minerals, Metals & Materials Society
ORIGINAL RESEARCH ARTICLE
Assessment of Deoxyribonuclease Activity Using DNA Molecules Immobilized Between Microelectrodes TAKAHIRO HIMURO
1,2
and YOJI SAITO1
1.—Department of Systems Design Engineering, Faculty of Science and Technology, Seikei University, 3-3-1 Kichijoji-Kitamachi, Musashino-shi, Tokyo 180-8633, Japan. 2.—e-mail: [email protected]
Serum deoxyribonuclease I (DNase I) can serve as a functional biomarker for the therapeutic monitoring of acute myocardial infarction and other diseases. Here, we demonstrate that the electrical properties of DNA molecules can be exploited to monitor enzymatic activity. A label-free DNA biosensor for the detection of DNase I activity was devised based on electrochemical impedance spectroscopy (EIS). Multiple lambda phage DNA molecules were immobilized between two electrodes in a polydimethylsiloxane reservoir. An equivalent circuit estimated from the EIS measurement was used to calculate the impedance of DNA molecules between the electrodes. DNase detection was then achieved by measuring the increase in impedance, after DNA cleavage by DNase I. This was assessed by the impedance-increase ratio, defined as Rafter/ Rbefore (where Rbefore and Rafter represent the resistance between the electrodeimmobilized DNA molecules before and after DNase I treatment, respectively). After treatment with DNase I at a concentration 102 unit/lL, a reproducible impedance-increase ratio of approximately 3.3 times was obtained, with a standard deviation of less than 20%. When DNase solutions of various concentrations were introduced, we succeeded in obtaining a definite correlation between DNase concentration and impedance-increase rate, within the range of 104 unit/lL to 101 unit/lL. Key words: DNA, DNase, impedance, equivalent circuit, EIS measurement
INTRODUCTION Deoxyribonuclease I (DNase I), which nonspecifically cleaves phosphodiester linkages of DNA, is implicated in several diseases.1–4 It is also known that DNase I activity in the serum is higher in patients with breast5,6 and oral cavity cancers.7 Furthermore, patients with prostate cancer8 and systemic lupus erythematosus (SLE),9,10 exhibit reduced DNase I activity, while those suffering from acute myocardial infarction (AMI) show increased DNase I activity.11,12 Consequently, the measurement of DNase I activity might be regarded as an innovative diagnostic marker for the early
(Received March 27, 2020; accepted October 30, 2020)
diagnosis of these syndromes. Moreover, genetic research involving DNA recombination shows that DNase I contamination of solutions causes undesired DNA cleavage, contributing to experimental failures. Therefore, sensitive detection of DNase is essential for success in genetic research. Several methods are available to assay DNase I activity. The fluorometric analyses13–15 generally used for DNase detection are useful, but have certain drawbacks, such as difficulty in automatic assay readout, time-co
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