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Microchimica Acta

, 186:158 | Cite as

Determination of Alzheimer biomarker DNA by using an electrode modified with in-situ precipitated molybdophosphate catalyzed by alkaline phosphatase-encapsulated DNA hydrogel and target recycling amplification

  • Xiaoyu Hua
  • Xingxing Zhou
  • Shijing Guo
  • Ting Zheng
  • Ruo Yuan
  • Wenju XuEmail author
Original Paper
  • 24 Downloads

Abstract

An electrochemical biosensor is described for highly sensitive determination of tDNA, an Alzheimer’s disease (AD)-related biomarker. Electroactive molybdophosphate anions were precipitated in-situ on a glassy carbon electrode (GCE) via catalytic hydrolysis by alkaline phosphatase (ALP). This is followed by recycling amplification of tDNA. Four DNA strands (referred to as S1, S2, S3 and S4) were designed to assemble X-shape DNA (X-DNA) building blocks. These were further extended into four directions under the action of DNA polymerase. The resultant two X-DNA motifs were polymerize. Simultaneously, ALP is encapsulated into a hydrogels network to obtain a porous material of type ALP@DNAhg. The GCE was modified with reduced graphene oxide functionalized with gold nanoparticles (Au@rGO). If ALP@DNAhg are captured via strand displacement, tDNA recycling assembly for signal amplification is initiated. This results in the immobilization of large amounts of ALP. On introduction of pyrophosphate and molybdate (MoO42−), ALP will catalyze the hydrolysis of pyrophosphate to produce phosphate. It will react with molybdate to form redox active phosphomolybdate anions (PMo12O403−). Its amperometrical signal depends on the concentration of tDNA in the 1.0 × 10−2 to 1.0 × 104 pM concentration range, and the detection limit is 3.4 × 10−3 pM.

Graphical abstract

Schematic presentation of (a) preparation of alkaline phosphatase-encapsulated DNA hydrogel (ALP@DNAhg). (b) fabrication of the biosensor for target DNA (tDNA) based on ALP@DNAhg to catalyze in situ precipitation of electroactive molybdophosphate anion (PMo12O403−) and tDNA recycling amplification, achieving tDNA-dependent electrochemical signal readout (X-DNA: X-shape DNA building block. TdT: terminal deoxynucleotidyl transferase. dATP: deoxyadenosine triphosphate. dTTP: deoxythymidine triphosphate. X-DNA-pAn and X-DNA-pTn: X-DNA motifs with poly-A and poly-T tails. ALP: alkaline phosphatase. ALP@DNAhg: ALP-encapsulated DNA hydrogels. Au@rGO: gold nanoparticles-functionalized reduced graphene oxide. GCE: glass carbon electrode. HP1, 2: hairpin DNA 1, 2. MCH: 6-mercaptohexanol. tDNA: target DNA. CV: cyclic voltammetry).

Keywords

Label-free detection Electrochemical biosensor AuNP-functionalized reduced graphene oxide X-shape DNA building block Pyrophosphate Molybdate In-situ precipitation Networked DNA structure Enzymatic catalysis Redox active phosphomolybdate anions 

Notes

Acknowledgements

The financial support by the National Natural Science Foundation (NNSF) of China (21775123) and the Natural Science Foundation Project of Chongqing (cstc2018jcyjAX0214) to this work was deeply appreciated.

Compliance with ethical standards

The author(s) declare that they have no competing interests.

Supplementary material

604_2019_3283_MOESM1_ESM.doc (540 kb)
ESM 1 (DOC 540 kb)

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Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Key Laboratory of Luminescent and Real-Time Analytical Chemistry, Ministry of Education, School of Chemistry and Chemical EngineeringSouthwest UniversityChongqingPeople’s Republic of China

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