A label-free aptasensor for highly sensitive detection of ATP and thrombin based on metal-enhanced PicoGreen fluorescence

Kaiyu Wang; Jian Liao; Xiangyue Yang; Meng Zhao; Min Chen; Weirong Yao; Weihong Tan; Xiaopeng Lan

doi:10.1016/j.bios.2014.07.022

A label-free aptasensor for highly sensitive detection of ATP and thrombin based on metal-enhanced PicoGreen fluorescence

Biosens Bioelectron. 2015 Jan 15:63:172-177. doi: 10.1016/j.bios.2014.07.022. Epub 2014 Jul 14.

Authors

Kaiyu Wang¹, Jian Liao², Xiangyue Yang², Meng Zhao², Min Chen², Weirong Yao³, Weihong Tan⁴, Xiaopeng Lan⁵

Affiliations

¹ Institute of Laboratory Medicine, Fuzhou General Hospital of Nanjing Military Command, Fuzhou 350025, PR China; Department of Laboratory Medicine, Dongfang Hospital, Xiamen University, Fuzhou 350025, PR China; Department of Laboratory Medicine, Fuzong Clinical College, Fujian Medical University, Fuzhou 350025, PR China.
² Institute of Laboratory Medicine, Fuzhou General Hospital of Nanjing Military Command, Fuzhou 350025, PR China; Department of Laboratory Medicine, Dongfang Hospital, Xiamen University, Fuzhou 350025, PR China.
³ Department of Laboratory Medicine, Fuzong Clinical College, Fujian Medical University, Fuzhou 350025, PR China.
⁴ Center for Research at the Bio/Nano Interface, Department of Chemistry and Shands Cancer Center, University of Florida, Gainesville, Florida 32611, United States; Plant Pathology Lab, North Florida Research and Education Center, University of Florida, Quincy, Florida 32351, United States; Molecular Science and Biomedicine Laboratory, State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Collaborative Innovation Center for Chemistry and Molecular Medicine, Hunan University, Changsha 410082, PR China. Electronic address: tan@chem.ufl.edu.
⁵ Institute of Laboratory Medicine, Fuzhou General Hospital of Nanjing Military Command, Fuzhou 350025, PR China; Department of Laboratory Medicine, Dongfang Hospital, Xiamen University, Fuzhou 350025, PR China; Department of Laboratory Medicine, Fuzong Clinical College, Fujian Medical University, Fuzhou 350025, PR China. Electronic address: lanxp@sina.com.

PMID: 25086329
DOI: 10.1016/j.bios.2014.07.022

Abstract

A label-free fluorescence aptasensor for highly selective and sensitive detection of ATP and thrombin was developed by using PicoGreen (PG) as signal molecule and surface-bound metal-enhanced fluorescence (MEF) substrates (silver island films, SIFs) as signal enhancers. On binding with ATP or thrombin, aptamers undergo structure switching, leading to a reduction of fluorescence intensity of PG. Chang of fluorescence intensity can be magnified by SIFs. The limit of detection for ATP and thrombin is 1.3 nM and 0.073 nM, respectively. The fluorescence quenching efficiency is linear in the logarithmic scale with ATP concentration range from 10 nM to 100 μM (R(2)=0.995) and thrombin concentration range from 0.1 nM to 100 nM (R(2)=0.997). The coefficients of variation of the intra-assay reproducibility and inter-assay reproducibility for ATP (10 μM) assay are 3.8% and 5.2%, respectively. In addition, the aptasensor is stable and can be reliably used for ATP measurement in biological samples. Overall, the aptasensor can be a useful and cost effective tool for the specific detection of ATP, thrombin and potentially other biomolecules in biological samples.

Keywords: ATP; Aptasensor; Label-free; Metal-enhanced fluorescence; PicoGreen; Thrombin.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adenosine Triphosphate / analysis*
Aptamers, Nucleotide / chemistry
Aptamers, Nucleotide / genetics*
Biosensing Techniques / instrumentation*
Complex Mixtures / analysis
Equipment Design
Equipment Failure Analysis
Fluorescent Dyes / analysis
Fluorescent Dyes / chemistry
Organic Chemicals / analysis
Organic Chemicals / chemistry
Reproducibility of Results
Sensitivity and Specificity
Silver / chemistry
Spectrometry, Fluorescence / instrumentation*
Spectrometry, Fluorescence / methods
Staining and Labeling
Thrombin / analysis*

Substances

Aptamers, Nucleotide
Complex Mixtures
Fluorescent Dyes
Organic Chemicals
PicoGreen
Silver
Adenosine Triphosphate
Thrombin