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ARTICLE IN PRESS
SNB-23328; No. of Pages 7
Sensors and Actuators B xxx (2017) xxx–xxx
Contents lists available at ScienceDirect
Sensors and Actuators B: Chemical
journal homepage: www.elsevier.com/locate/snb
Construction of a paper-based electrochemical biosensing platform
for rapid and accurate detection of adenosine triphosphate (ATP)
Po Wang a,b,∗ , Zhiyuan Cheng a , Qian Chen a , Lulu Qu a , Xiangmin Miao a , Qiumei Feng a
a
b
School of Chemistry and Materials Science, Jiangsu Normal University, Xuzhou 221116, China
Department of Chemistry and Biochemistry, Florida International University, Miami 33199, United States
a r t i c l e
i n f o
Article history:
Received 15 April 2017
Received in revised form 2 October 2017
Accepted 3 October 2017
Available online xxx
Keywords:
ATP
Electrochemical detection
Paper-based electrode
Biosensing platform
a b s t r a c t
In this paper, a simple but efficient electrochemical biosensing platform was developed for ATP assay
based on the preparation of paper-based working electrodes. In contrast to the electrochemical response
of commercial glassy carbon electrode (GCE), improved voltammetric signals were achieved at the paperbased electrode. In particular, the surface contamination arising from oxidation product, a prominent
challenge for ATP detection, was effectively eliminated by the paper-based system, endowing reliable
analysis with high reproducibility. Moreover, the practical application value of the system was verified
by assay of ATP in human serums, cancer cells, and normal cells with satisfactory results. Compared
with the detection performances of traditional strategies, the advantages involved in the system were
exhibited to be rapid, accurate, convenient, and inexpensive, which confer the proposed sensing platform
promising for applications in public health as well as the fundamental research of molecular biology.
© 2017 Elsevier B.V. All rights reserved.
1. Introduction
Adenosine triphosphate (ATP), the primary energy currency
in living organisms, plays vital roles in biosynthesis, cellular
metabolism, DNA replication and transcription, and the regulation of biochemical pathways in cell physiology [1]. Besides, ATP
is identified to act as a signaling agent in the cascade for the
modulation of peripheral and central nervous system [2]. Further studies revealed that ATP can be used as an indicator for
cell injury, viability, and proliferation [3]. Abnormal level of ATP
in organism is closely related to various pathogenesis, including
hypoxia, ischemia, hypoglycemia, Alzheimer’s disease, and Parkinson’s disease [2]. In recent years, a variety of intriguing approaches
have been developed to detect ATP, such as photoelectrochemical
aptasensor [4], surface-enhanced Raman scattering [5], optomagnetic aptasensor [6], electrochemiluminescence [7], fluorescent
spectroscopy [8], and chemiluminescence resonance energy transfer [9]. These methods display apparent merits for ATP detection,
however, complicated operations, insufficient sensitivities, expensive equipments, or large background interferences are commonly
involved [4,10], which limit their potential application fields.
∗ Corresponding author at: School of Chemistry and Materials Science, Jiangsu
Normal University, Xuzhou 221116, China.
E-mail address: wangpo@jsnu.edu.cn (P. Wang).
Electrochemical technique is proved to be a powerful tool for
ATP analysis due to its inherent advantages of low cost, high sensitivity and selectivity, and ease of operation [11]. In particular,
electrochemical aptamer-based biosensor has attracted increasing attention owing to the high binding affinity and specificity, as
well as facile combination with amplification approaches [4,11].
In 2007, Zuo et al. reported a target-responsive electrochemical
aptamer switch, which pioneered a conceptually new approach for
reagentless assay of ATP [12]. Afterward, a multifunctional labelfree electrochemical biosensor was developed for parallel detection
of ATP and thrombin based on an integrated aptamer [13]. According to the ingenious design of single aptamer sequences, Plaxco’s
group proposed an innovative sandwich detection approach for
small-molecule targets of ATP and cocaine [14]. Interestingly, an
autonomous bio-barcode DNA machine combined with an aptamer
recognition element was created for determination of ATP in tumor
cells [15]. Taking advantage of nanocomposite modified carbon
paste electrode, real-time monitoring of ATP was realized in a
wide concentration range [16]. In order to improve the specificity for target analysis, a dual recognition unit strategy was
designed for cerebral ATP assay [17]. Moreover, some signal amplification strategies were developed to improve the sensitivity for
ATP detection, which were based on quantum dots [18], functionalized gold nanoparticles [19], rolling circle amplification [20],
nuclease-assisted target recycling [21], as well as hybridization
chain reaction-induced DNA concatamer [22].
https://doi.org/10.1016/j.snb.2017.10.024
0925-4005/© 2017 Elsevier B.V. All rights reserved.
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
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2
Electrochemical aptasensors confer the analysis of ATP with
high sensitivity and selectivity, while the detection procedures are
associated with time-intensive incubation, conjugation or enzymatic reaction steps [12,16,19], which inevitably decrease the
analysis speed. Compared with the detection performances of
aptasensors, the methodology based on the direct electrochemical
oxidation of analyte is a more straightforward, rapid and convenient approach [23,24]. Taking advantage of the electroactive
moiety of adenine, the oxidation signal of adenosine was successfully acquired at a nanocarbon film electrode in Niwa’s group
[23,25]. Based on the preparation of carboxyl functionalized ionic
liquid modified carbon paste electrode, the oxidation current of
ATP was measured for quantitative determination [26]. Besides,
boron-doped diamond electrode was demonstrated to be an effective electrochemical interface for adenosine detection [27,28].
However, a significant challenge was encountered for direct electrochemical sensing of ATP because of the fouling effect resulted
from the adsorption of oxidation product. The electrochemical
oxidation of purine and its derivatives is irreversible, including adenine, adenosine, and ATP [25]. It was revealed that the reaction
product of adenine irreversibly adsorbed on the sensing interface,
which passivated the electroactive of conventional working electrodes [29], leading to poor sensitivity and reproducibility.
Inspired by Martinez et al. from Harvard University [30], paperbased analytical devices (PADs) pave a revolutionary route for
exploring electrochemical and biological analysis methodologies.
Herein, a simple but efficient electrochemical biosensing platform
was developed for ATP detection based on the preparation of
paper-based electrodes using vacuum filtration system. In contrast
to traditional working electrodes, the paper-based electrodes are
extremely inexpensive, which can be used as disposable sensing
device for target analysis. As a result, the fouling effect arising from
the accumulation of oxidation product on the sensing interface was
effectively eliminated without the need for additional treatment.
In comparison with the electrochemical response of ATP at commercial glassy carbon electrode (GCE), higher current sensitivity
and lower overpotential were exhibited at the paper-based device.
Moreover, the proposed sensing platform realized the assay of ATP
in human serums, cancer cells, and normal cells in an accurate,
rapid, convenient, and reproducible way, confirming the practical
application ability of the system in public health and biochemical
studies.
2. Experimental
2.1. Materials and reagents
ATP was purchased from Sigma-Aldrich (USA). Mixed cellulose ester (MCE) filter paper with 100 nm pore size was obtained
from Merck Millipore (Darmstadt, Germany). Fast drying silver
paint was purchased from Ted Pella Inc. (California, USA). Potassium ferricyanide (K3 [Fe(CN)6 ]) and graphite (spectral pure) were
obtained from Sinopharm Chemical Reagent Co., Ltd. (Shanghai,
China). Acetate buffers with different pH were prepared by mixing stock solutions of 0.1 M HAc and NaAc. All the chemicals with
analytical reagent grade were used without further purification.
Deionized water (specific resistance >18.2 M cm−1 ) was obtained
from a Milli-Q gradient system (Millipore) for the preparation of
aqueous solutions.
2.2. Apparatus and measurements
The vacuum filtration operations for the preparation of paperbased electrodes were carried out on a KONTES Ultra-Ware system
(Kimble Chase, Vineland, USA). Differential pulse voltammetry
(DPV) and cyclic voltammetry (CV) were performed on a CHI660E
electrochemical workstation (CH Instruments, Austin, USA). The
three-electrode system consisted of a paper-based working electrode, a platinum wire counter electrode, and an Ag/AgCl wire
reference electrode. Before electrochemical measurements, ultrapure nitrogen was used to eliminate the interference from the
reduction of oxygen in electrolyte solutions. Field emission scanning electron microscope (FE-SEM) images were acquired on an
S-4800 microanalyzer (Hitachi, Japan). X-ray photoelectron spectroscopy (XPS) was carried out on an ESCALAB 250 spectrometer
equipped with a monochromatic Al K␣ X-ray source (Thermo Fisher
Scientific Inc., U.K.). Raman spectra were recorded on a LabRAM
HR800 spectrometer at an excitation wavelength of 532 nm (Horiba
Jobin Yvon, France). High performance liquid chromatography
(HPLC) measurements were performed on an Agilent 1100 Series
chromatograph equipped with a quaternary pump, a diode array
detector (DAD), and an autosampler (Agilent Technologies, California, USA).
2.3. Preparation of paper-based electrodes
For the preparation of graphene-functionalized paper chips,
graphene oxide was synthesized from graphite by an improved
Hummers method [31], followed by the chemical reduction with
hydrazine [32]. Reduced graphene oxide was dispersed in deionized water with the treatment of sonication for 20 min, resulting
in a 6.0 mg L−1 homogeneous dispersion. The paper-based chips
were prepared according to Wu’s work with modifications and
improvements [33,34]. In a typical procedure, the MCE membrane
was firstly cleaned with 20 mL deionized water for three times on
vacuum filtration system. Then, 15 mL of the prepared graphene
suspension was cautiously loaded onto the cleaned MCE filter
paper. After immersion for 5 min, the vacuum filtration operations
were carried out to form a uniform graphene assembled film. Subsequently, the graphene-functionalized MCE was rinsed with water
for six times to get rid of foreign substances. Afterward, the resulting film was vacuum-dried overnight in nitrogen atmosphere at
ambient temperature. In order to prepare paper-based working
electrodes, the vacuum-dried film was carefully transferred onto a
cutting template, and cut into 18 independent chips. Taking advantage of fast drying silver paint, these chips were connected to
copper wires, and then wrapped with Parafilm to form working
electrodes (graphene/MCE). In control experiments, commercial
glassy carbon electrode (GCE) was sequentially polished with 1.0,
0.3, and 0.05 ␮m Al2 O3 slurries, and cleaned with sonication for
10 min before use.
2.4. Cellular ATP analysis
The assay of cellular ATP was implemented in K562 leukemia
cells, human adenocarcinoma HeLa cells, and human normal breast
cells (MCF-10A). The cells were separately cultured in Dulbecco’s
modified Eagle medium (DMEM) supplemented with 10% fetal
bovine serum (FBS) and 5% humidified CO2 at 37 ◦ C, and the cells
were harvested by trypsinization. The cell density was determined
by a hemocytometer prior to each experiment. Then, the suspensions of K562 cells, HeLa cells, and MCF-10A cells, dispersed in
DMEM cell media buffer, were centrifuged at 3000 rpm for 5 min,
and washed with phosphate-buffered saline (18.6 mM phosphate,
4.2 mM KCl, and 154.0 mM NaCl, pH 7.4) five times and resuspended in deionized water. Finally, the cells were disrupted by
sonication for 20 min at 0 ◦ C, and the resulting lysates were centrifuged at 12000 rpm for 15 min at 4 ◦ C to remove the homogenate
of cell debris. The supernatants were collected and deproteinized
through Amicon YM10 membrane (Millipore Corp., Bedford, MA).
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
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The freshly extracted cell lysates were diluted with acetate buffer
before electrochemical detection.
3. Results and discussion
3.1. The interfacial structure and property of the sensing system
FE-SEM was employed to study the interfacial structure of
the paper-based system. Fig. 1A described the typical framework
of MCE membrane, which exhibited porous nanonetwork morphology. The MCE nanowires were intertwined together to form
a permeable structure with the nanopore diameter of ∼100 nm.
Taking advantage of the vacuum filtration technique, graphene
nanosheets were compactly and homogeneously assembled onto
the surface of MCE paper (Fig. 1B). The typically wrinkled structure
of graphene nanosheets with average size of 3–5 ␮m and thickness
of 1–3 nm was clearly observed. The resulting graphene film significantly increased the roughness and electroactive surface area
of the sensing interface, which were anticipant for electrochemical
applications. Moreover, the element and composition informations
of the resulting graphene film were confirmed by XPS characterization. As shown in Fig. 1C, the predominant C1 s and O1 s signal peaks
were clearly displayed at 285.0 and 531.1 eV, respectively, while
almost no other peak related to external contamination could be
seen. Quantitative peak deconvolution of the XPS survey rendered
a 100:9.3 atomic ratio of C/O, which was close to the reported value
[32]. The chemical structure of the prepared reduced graphene
oxide was further investigated by Raman spectroscopy. As shown
in Fig. 1D, only G band could be observed for graphite (curve a),
while both D and G bands were clearly presented at 1349 and
1594 cm−1 for reduced graphene oxide (curve b), corresponding
to sp3 -hybridized carbon and E2g zone center mode of the crystalline graphite, respectively [35]. It was revealed that the relative
intensity of the D and G lines (ID /IG ratio) was closely related to the
amount of defect sites in carbonaceous materials [35]. Compared
with graphite, the ID /IG ratio of reduced graphene oxide increased
remarkably, indicating that there were significant edge-plane-like
defective sites existing on the surface of reduced graphene oxide
[32], which were favorable for the electrochemical oxidation of
ATP. In addition, the electrochemical property of the proposed
graphene/MCE was comparatively investigated with commercial
GCE using redox probe [Fe(CN)6 ]3− (data not shown). The results
demonstrated that a pair of quasi-reversible redox peaks with a
peak separation (Ep ) of 88 mV was presented at the commercial
GCE. In contrast, the peak currents increased distinctly and the Ep
reduced to 71 mV at the graphene/MCE, revealing that the paperbased electrode facilitated the electron transfer rate constant of
[Fe(CN)6 ]3−/4− redox reaction.
3.2. Direct oxidation behavior of ATP
As shown in Fig. 2A, the electrochemical oxidation of ATP was
explored at commercial GCE, graphene/MCE and graphene/GCE in
acetate buffer. It was demonstrated that a very weak and broad oxidation peak was displayed at 1.41 V for commercial GCE (curve a,
green), corresponding to the slow electron transfer behavior of GCE.
In the case of graphene/MCE, the peak current increased greatly, the
peak width at half height (W1/2 ) decreased apparently, and the peak
potential shifted negatively to 1.34 V (curve b, blue), which indicated that graphene film effectively promoted the electron transfer
kinetics involved in the oxidation reaction of ATP. The catalytic
mechanism was attributed to the fact that the graphene nanosheets
markedly increased the edge-plane-like defective site density of the
electrochemical reaction interface. Moreover, control experiment
was performed at graphene/GCE for ATP detection. Compared with
3
the electrochemical response of graphene/MCE (curve b, blue), relatively lower current sensitivity as well as higher overpotential was
presented at graphene/GCE (curve c, red), revealing that the paperbased electrode provided a more suitable electrochemical interface
for ATP detection. It should be noted that ATP was oxidized at relatively higher potential (more than 1.3 V). The application of higher
potential resulted in larger background signal of blank solution.
Consequently, the peak current of ATP was masked partially, which
disturbed the signal recognition. In order to overcome this problem,
blank experiments were performed in parallel at the same working conditions, and the contributions of the background signals of
blank solution were presented in Fig. 2A (black curves). As a result,
the oxidation currents of ATP could be discriminated distinctly by
subtracting the background currents of blank solution. After the
subtraction, a more clear resolution of oxidation signal of ATP was
achieved (Fig. 2B).
3.3. Effect of pH on the voltammetric response of ATP
The influence of solution pH on the electrochemical oxidation
of ATP was investigated in acetate buffer. As presented in Fig. 3A,
the peak potential (Epa ) for DPV response of ATP moved to positive direction with the decrease in solution pH from 9.0 to 3.0,
which suggested that the oxidation reaction of ATP was associated
with proton-transfer process. The relationship between Epa and pH
was depicted as Epa (V) = 1.609–0.0556 pH (n = 6, R = 0.9991). The
resulting slope of 55.6 mV pH−1 was close to the theoretical value
of 59.1 mV pH−1 at 25 ◦ C, indicating that the electrochemical oxidation of ATP involved an equal number of protons and electrons. The
oxidation mechanism of ATP was proposed in Fig. 3B, which was
consistent with literature reports [26,36]. With regard to the effect
of solution pH on the current response of ATP, it was observed that
the maximum current density appeared at pH 5.0. Since the pKa
of HAc is 4.75, in order to keep the maximum buffer capacity for
real sample analysis, pH 4.75 acetate buffer was selected as the
supporting electrolyte in this work.
3.4. Optimization of the amount of graphene for electrode
preparation
The condition experiment for optimizing the amount of
graphene on the MCE was carried out. The results displayed that the
current density of ATP increased significantly with the increase in
the volume of graphene suspension (6.0 mg L−1 ) from 1.0 to 12 mL,
which was attributed to the improvement of the conductivity and
electroactive surface area of the resulting sensing interface. However, a relatively stable current response was obtained when the
volume of graphene suspension was more than 15 mL, suggesting that the MCE paper was assembled with sufficient graphene
nanosheets for ATP detection, and the physicochemical properties of the resulting graphene/MCE were good enough for sensing
applications. Therefore, 15 mL graphene suspension was chosen as
the optimal condition for electrode preparation, and the absolute
amount of graphene on MCE was calculated to be 90 ␮g.
3.5. Quantitative analysis of ATP
The disposable graphene/MCE provided a powerful tool for
accurate detection of ATP without fouling effect. As described in
Fig. 4A, with the gradual increase in ATP concentration, the peak
potential was constantly located at 1.34 V, illustrating the stable
and reliable electrochemical response of the paper-based sensing
system. The current density of ATP was proportional to the concentration in the range of 0.3–450 ␮M, with the linear regression
equation of jpa (␮A cm−2 ) = 2.598c (␮M) + 1.275 (n = 10, R = 0.9985).
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
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Fig. 1. (A) FE-SEM image of the MCE membrane. (B) FE-SEM image of the graphene/MCE chip. (C) XPS spectrum of the graphene functionalized MCE. (D) Raman spectra of
graphite (a) and reduced graphene oxide (b).
Fig. 2. (A) DPVs of 50 ␮M ATP (colour curves) and blank solution (black curves) at commercial GCE (a), graphene/MCE (b) and graphene/GCE (c) in 0.1 M pH 4.75 acetate
buffer. (B) Background-subtracted DPVs of 50 ␮M ATP at commercial GCE (a), graphene/MCE (b) and graphene/GCE (c) in 0.1 M pH 4.75 acetate buffer.
Fig. 3. (A) Background-subtracted DPVs of 50 ␮M ATP at graphene/MCE in 0.1 M acetate buffer with different pH (from a to g: 9.0, 8.0, 7.0, 6.0, 5.0, 4.0, 3.0). (B) Electrochemical
reaction mechanism of ATP at the paper-based sensing system.
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
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Fig. 4. (A) Background-subtracted DPVs of graphene/MCE in 0.1 M pH 4.75 acetate buffer containing ATP with concentrations of 2, 5, 10, 15, 25, 35, 50, 70, 90, and 120 ␮M
(from 1 to 10, respectively). (B) The linear relationship between current densities and concentrations.
The calibration curve was shown in Fig. 4B, and the limit of detection (LOD) was estimated to be 0.08 ␮M (S/N = 3).
3.6. Interference investigations
The interferential behavior of foreign species on ATP detection was studied by adding various interferents into 50 ␮M ATP
for electrochemical detection. It was demonstrated that common
metal ions and acid radical ions exhibited almost no interference
in a 100-fold concentration (signal change <1%), such as K+ , Ca2+ ,
Na+ , Zn2+ , Mg2+ , Fe3+ , CO3 2− , HCO3 − , Cl− , PO4 3− , SO4 2− and NO3 − .
The influences of some redox active substances which may exist
in biological samples were evaluated, such as dopamine, uric acid,
ascorbic acid, glucose, hemoglobin, norepinephrine and serotonin.
Although these electroactive molecules could generate oxidation
currents at the sensing interface, their peak potentials were much
negative compared with that of ATP (1.34 V). The results revealed
that the current changes of ATP were less than 5% of the initial
signal after the addition of the above substances in a 50-fold content. Moreover, the effect of purine base-containing substances
on ATP detection was also examined, such as guanine, adenine,
adenosine, AMP, ADP and dATP. It was found that the oxidation
signals of guanine and adenine were clearly separated from that
of ATP with sufficient potential resolutions, and the peak potentials of adenosine, AMP, ADP and dATP were close to 1.34 V (ATP).
Therefore, pretreatment step or mathematical deconvolution technique is required for the analysis of mixed ATP, AMP, ADP, dATP and
adenosine.
3.7. Stability and reproducibility of the system
Stability and reproducibility are important criterions for evaluating the detection performance of a sensing platform. The stability
of the proposed system was tested by periodical detection of 50 ␮M
ATP. It was demonstrated that the current signal of graphene/MCE
retained 97.2% of the initial voltammetric response after one week
storage at room temperature. The signal decline was less than 8%
after one month, indicating that the paper-based electrodes can
be used for persistent operation. It should be noted that no significant change in current response was detected after storage at
95 ◦ C for 24 h (signal change < 5%), which was ascribed to the inherent thermal stability of graphene material. The reproducibility of
the system was examined by successive detection of 50 ␮M ATP
using 12 electrodes. The results showed that the relative standard deviation (R.S.D.) of the current densities for 12 independent
measurements was calculated to be 3.9%, revealing an acceptable
precision for ATP assay.
3.8. Specificity of the sensing system
Specificity is an important factor for evaluating the detection performance of a sensing system, especially for potential
applications in complicated biological samples. Belonging to
the nucleoside triphosphate family, cytidine triphosphate (CTP),
guanosine triphosphate (GTP), thymidine triphosphate (TTP) and
uridine triphosphate (UTP) usually coexist with ATP in real samples,
which were commonly used as controls for specificity investigation
[5,21,37,38]. In order to explore the specificity of the proposed system for target analysis, the paper-based electrode was challenged
with electrochemical detection of ATP analogues under the same
experimental conditions. The results demonstrated that the oxidation peak potentials of CTP, GTP, TTP and UTP were far from that of
ATP (1.34 V), and the corresponding current signals were negligible
at 1.34 V. Moreover, when 50 ␮M ATP was mixed with 10-fold concentration of CTP, GTP, TTP and UTP, the resulting signal change
at 1.34 V was less than 5% of the current response of ATP alone,
indicating acceptable specificity of the system for ATP analysis.
3.9. Sensing applications for real samples
The proposed system was challenged with the assay of ATP in
human whole blood serums to examine the validity of the platform.
The serum samples donated by healthy volunteers were collected
from Xuzhou central hospital in Jiangsu Province, China. Since the
oxidation peak potential of ADP was close to that of ATP, deconvolution technique was applied for their signal discrimination. Based
on the separated current signals of ATP, the detection results of four
different samples were listed in Table 1, which were close to literature values [39]. It was reported that the concentration of ATP was
about 2–4 folds of ADP in blood samples [40,41], which indicated
the approximate content level of ADP in these samples. The reliability of the system was verified by HPLC determinations of the
same samples. As described in Table 1, the detection results of the
two methods were in good agreement for sample analysis. Therefore, the paper-based system provided an efficient platform for the
assay of ATP in real biological samples.
The practical application performance of the paper-based sensing platform was further confirmed by detection of ATP in cancer
cells and normal cells, including K562 leukemia cells, human adenocarcinoma HeLa cells, and human normal breast cells (MCF-10A).
The cell lysates were freshly extracted, and were subsequently
diluted with acetate buffer prior to analysis. As shown in Table 2, the
cellular ATP levels for K562, HeLa and MCF-10A cells were obtained
to be 2.458, 2.163 and 1.097 mM, respectively, which were consistent with the ATP concentrations that are typically found in cells
[42]. After adding 1.000, 2.000 and 3.000 mM ATP into the sam-
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
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Table 1
Detection results of ATP in human whole blood serum samples (n = 6).
Sample
1
2
3
4
Paper-based sensing platform
HPLC method Detected (␮M)
Detected (␮M)
Added (␮M)
Found (␮M)
Recovery (%)
R.S.D. (%)
0.896
0.945
1.138
0.977
1.000
2.000
3.000
5.000
1.935
2.911
4.024
6.117
103.9
98.3
96.2
102.8
4.5
3.6
3.9
4.2
0.861
0.974
1.092
0.935
Table 2
Detection results of ATP in K562 cells, HeLa cells, and normal MCF-10A cells (n = 6).
Sample
K562
HeLa
MCF-10A
Paper-based sensing platform
HPLC method Detected (mM)
Detected (mM)
Added (mM)
Found (mM)
Recovery (%)
R.S.D. (%)
2.458
2.163
1.097
1.000
2.000
3.000
3.412
4.105
4.223
95.4
97.1
104.2
3.8
4.9
4.0
2.360
2.072
1.135
Table 3
Comparison of analytical parameters of different detection methods for the assay of ATP.
Detection method
Linear range
Detection limit
Analysis time
Reference
Photoelectrochemical aptasensor
Surface-enhanced Raman scattering
Optomagnetic biosensor
Electrochemiluminescence
Fluorescent spectroscopy
Chemiluminescence
Photoluminescent sensing probe
Quartz crystal microbalance
Bioluminescent detection
Colorimetric analysis
Electrochemical oxidation a
Electrochemical oxidation b
Electrochemical oxidation c
Electrochemical oxidation d
Electrochemical oxidation
0.5 pM–5 nM
0.1–100 ␮M
100–10000 ␮M
0.1–1.0 ␮M
5–250 ␮M
100–1200 ␮M
42–324 ␮M
2–10 nM
5–100 ␮M
0.04–0.4 ␮M
5–1000 ␮M
20–200 ␮M
10–500 ␮M
0.10–10.0 ␮M
0.3–450 ␮M
0.18 pM
0.02 ␮M
74 ␮M
0.1 ␮M
5 ␮M
10 ␮M
19 ␮M
1.3 nM
1 ␮M
0.04 ␮M
1.67 ␮M
2 ␮M
1 ␮M
0.10 ␮M
0.08 ␮M
>18 h
>4 h
>80 min
>2.5 h
>65 min
>3 h
>70 min
>4 h
>24 h
>3 h
∼10 min
∼30 min
∼10 min
∼10 min
∼10 min
[4]
[5]
[6]
[7]
[8]
[9]
[43]
[44]
[45]
[46]
[26]
[27]
[47]
[48]
This work
a
b
c
d
Electrochemical oxidation based on ionic liquid modified carbon paste electrode.
Electrochemical oxidation based on cathodically treated boron-doped diamond electrode.
Electrochemical oxidation based on pencil graphite carbon electrode.
Electrochemical oxidation based on nanogold modified indium tin oxide electrode.
ples, the recoveries were calculated to be 95.4%, 97.1% and 104.2%,
respectively, with R.S.D. in the range of 3.8–4.9%. In addition, the
cellular ATP levels were further detected by HPLC method, which
validated the reliability of the proposed system for sensing applications.
3.10. Detection performance of the system
The detection performance of the sensing system was compared
with literatures based on three important analytical parameters, including linear range, detection limit, and analysis time. As
described in Table 3, the linear range and detection limit of the
proposed sensor were comparable with most reference methods,
while the analysis speed of the present work was remarkably
faster than reference methods [4–9,43–46]. The performance of
rapid analysis was attributed to the intrinsic detection mechanism of the approach, which was based on the direct oxidation
behavior of adenine in ATP molecular structure without the need
for time-consuming incubation, conjugation or enzymatic reaction
steps. Moreover, the analytical parameters of some electrochemical sensors based on ATP oxidation were also listed in Table 3 for
comparison. It was found that the analysis speed of the paper-based
electrode was similar to those of conventional chemically modified
electrodes [26,27,47,48], and the proposed sensing system exhibited relatively wider linear range and lower detection limit for ATP
detection.
4. Conclusions
In summary, a rapid, accurate, convenient, and inexpensive
approach was described for ATP analysis based on the construction of paper-based electrochemical biosensing platform.
Taking advantage of the functionalization of graphene nanosheets,
the electrochemical signal of ATP was clearly acquired at the
paper-based system without the need for designing complicated
aptasensor, which significantly accelerated the analysis speed.
Compared with the electrochemical response of commercial GCE,
the paper-based electrode demonstrated improved voltammetric
signals for ATP detection, including higher current sensitivity and
lower overpotential. Particularly, the paper-based electrode was
a disposable detection device, which could effectively eliminate
the surface contamination resulted from the adsorption of reaction product. The present work implemented the assay of ATP in
real complex samples within 10 min with acceptable results, which
illuminated the practical application fields of the sensing platform.
Acknowledgements
This work was supported by the National Natural Science Foundation of China (No. 21675067, 21205052), the Qing Lan project of
Jiangsu Province for outstanding teachers, and the project funded
by the Priority Academic Program Development of Jiangsu Higher
Education Institutions.
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
G Model
SNB-23328; No. of Pages 7
ARTICLE IN PRESS
P. Wang et al. / Sensors and Actuators B xxx (2017) xxx–xxx
References
[1] P.B. Dennis, A. Jaeschke, M. Saitoh, B. Fowler, S.C. Kozma, G. Thomas, Science
294 (2001) 1102–1105.
[2] A.V. Gourine, E. Llaudet, N. Dale, K.M. Spyer, Nature 436 (2005) 108–111.
[3] J.A. Cruz-Aguado, Y. Chen, Z. Zhang, N.H. Elowe, M.A. Brook, J.D. Brennan, J.
Am. Chem. Soc. 126 (2004) 6878–6879.
[4] G.C. Fan, M. Zhao, H. Zhu, J.J. Shi, J.R. Zhang, J.J. Zhu, J. Phys. Chem. C 120
(2016) 15657–15665.
[5] S. Ye, Y. Wu, X. Zhai, B. Tang, Anal. Chem. 87 (2015) 8242–8249.
[6] J. Yang, M. Donolato, A. Pinto, F.G. Bosco, E.T. Hwu, C.H. Chen, T.S. Alstrøm,
G.H. Lee, T. Schäfer, P. Vavassori, A. Boisen, Q. Lin, M.F. Hansen, Biosens.
Bioelectron. 75 (2016) 396–403.
[7] L. Hu, Z. Bian, H. Li, S. Han, Y. Yuan, L. Gao, G. Xu, Anal. Chem. 81 (2009)
9807–9811.
[8] G. Garai-Ibabe, M. Möller, L. Saa, R. Grinyte, V. Pavlov, Anal. Chem. 86 (2014)
10059–10064.
[9] X. Liu, R. Freeman, E. Golub, I. Willner, ACS Nano 5 (2011) 7648–7655.
[10] X. Li, Y. Peng, Y. Chai, R. Yuan, Y. Xiang, Chem. Commun. 52 (2016) 3673–3676.
[11] M. Labib, E.H. Sargent, S.O. Kelley, Chem. Rev. 116 (2016) 9001–9090.
[12] X. Zuo, S. Song, J. Zhang, D. Pan, L. Wang, C. Fan, J. Am. Chem. Soc. 129 (2007)
1042–1043.
[13] Y. Du, B. Li, H. Wei, Y. Wang, E. Wang, Anal. Chem. 80 (2008) 5110–5117.
[14] X. Zuo, Y. Xiao, K.W. Plaxco, J. Am. Chem. Soc. 131 (2009) 6944–6945.
[15] H. Zhang, C. Fang, S. Zhang, Chem. Eur. J. 16 (2010) 12434–12439.
[16] B.J. Sanghavi, S. Sitaula, M.H. Griep, S.P. Karna, M.F. Ali, N.S. Swami, Anal.
Chem. 85 (2013) 8158–8165.
[17] P. Yu, X. He, L. Zhang, L. Mao, Anal. Chem. 87 (2015) 1373–1380.
[18] H. Zhang, B. Jiang, Y. Xiang, Y. Zhang, Y. Chai, R. Yuan, Anal. Chim. Acta 688
(2011) 99–103.
[19] T. Zhao, R. Liu, X. Ding, J. Zhao, H. Yu, L. Wang, Q. Xu, X. Wang, X. Lou, M. He, Y.
Xiao, Anal. Chem. 87 (2015) 7712–7719.
[20] Y. Guo, X. Sun, G. Yang, J. Liu, Chem. Commun. 50 (2014) 7659–7662.
[21] D. Tang, J. Tang, Q. Li, B. Su, G. Chen, Anal. Chem. 83 (2011) 7255–7259.
[22] B.C. Yin, Y.M. Guan, B.C. Ye, Chem. Commun. 48 (2012) 4208–4210.
[23] D. Kato, N. Sekioka, A. Ueda, R. Kurita, S. Hirono, K. Suzuki, O. Niwa, J. Am.
Chem. Soc. 130 (2008) 3716–3717.
[24] P. Wang, P. Han, L. Dong, X. Miao, Electrochem. Commun. 61 (2015) 36–39.
[25] T. Kamata, D. Kato, S. Hirono, O. Niwa, Anal. Chem. 85 (2013) 9845–9851.
[26] H. Gao, M. Xi, L. Xu, W. Sun, Microchim. Acta 174 (2011) 115–122.
[27] K. Asai, T.A. Ivandini, M.M. Falah, Y. Einaga, Electroanalysis 28 (2016) 177–182.
[28] J.A. Birbeck, T.A. Mathews, Anal. Chem. 85 (2013) 7398–7404.
[29] O. Akhavan, E. Ghaderi, R. Rahighi, ACS Nano 6 (2012) 2904–2916.
[30] A.W. Martinez, S.T. Phillips, M.J. Butte, G.M. Whitesides, Angew. Chem. Int. Ed.
46 (2007) 1318–1320.
[31] W.S. Hummers, R.E. Offeman, J. Am. Chem. Soc. 80 (1958) 1339.
[32] M. Zhou, Y. Zhai, S. Dong, Anal. Chem. 81 (2009) 5603–5613.
[33] Z. Wu, Z. Chen, X. Du, J.M. Logan, J. Sippel, M. Nikolou, K. Kamaras, J.R.
Reynolds, D.B. Tanner, A.F. Hebard, A.G. Rinzler, Science 305 (2004)
1273–1276.
[34] B. Guntupalli, P. Liang, J.H. Lee, Y. Yang, H. Yu, J. Canoura, J. He, W. Li, Y.
Weizmann, Y. Xiao, ACS Appl. Mater. Interfaces 7 (2015) 27049–27058.
[35]
[36]
[37]
[38]
[39]
[40]
[41]
[42]
[43]
[44]
[45]
[46]
[47]
[48]
7
A.C. Ferrari, J. Robertson, Phys. Rev. B 61 (2000) 14095–14107.
B.E.K. Swamy, B.J. Venton, Anal. Chem. 79 (2007) 744–750.
H. Xie, Y. Chai, Y. Yuan, R. Yuan, Chem. Commun. 53 (2017) 8368–8371.
X. Chen, L. Ge, B. Guo, M. Yan, N. Hao, L. Xu, Biosens. Bioelectron. 58 (2014)
48–56.
T.W. Traut, Mol. Cell. Biochem. 140 (1994) 1–22.
T.O. Kiviniemi, G.G. Yegutkin, J.O. Toikka, S. Paul, T. Aittokallio, T. Janatuinen, J.
Knuuti, T. Rönnemaa, J.W. Koskenvuo, J.J. Hartiala, S. Jalkanen, O.T. Raitakari,
Front. Physiol. 3 (2012) 1–8.
N. Bakhtiari, S. Hosseinkhani, B. Larijani, M.R. Mohajeri-Tehrani, A. Fallah, J.
Diabetes Metab. Dis. 11 (2012) 9.
D. Zheng, D.S. Seferos, D.A. Giljohann, P.C. Patel, C.A. Mirkin, Nano Lett. 9
(2009) 3258–3261.
K. Selvaprakash, Y.C. Chen, Biosens. Bioelectron. 61 (2014) 88–94.
W. Song, Z. Zhu, Y. Mao, S. Zhang, Biosens. Bioelectron. 53 (2014) 288–294.
K. Tsukagoshi, M. Tahira, R. Nakajima, J. Chromatogr. A 1094 (2005) 192–195.
L. Zhang, S. Guo, J. Zhu, Z. Zhou, T. Li, J. Li, S. Dong, E. Wang, Anal. Chem. 87
(2015) 11295–11300.
K. Kerman, M. Vestergaard, E. Tamiya, Anal. Lett. 41 (2008) 2077–2087.
R.N. Goyal, M. Oyama, S.P. Singh, Electroanalysis 19 (2007) 575–581.
Biographies
Po Wang obtained his Ph.D. degree in Analytical Chemistry from Sun Yat-Sen
University, China, in 2011. Then, he worked as an associate professor in Jiangsu Normal University. His scientific interests mainly include electroanalytical chemistry,
biosensors, and the synthesis and applications of nanomaterials.
Zhiyuan Cheng is now a graduate student in School of Chemistry and Materials
Science, Jiangsu Normal University. His research interests are mainly focus on electroanalytical chemistry and biosensors.
Qian Chen is now a graduate student in School of Chemistry and Materials Science,
Jiangsu Normal University. Her research interests are mainly focus on biosensors
and electroanalytical chemistry.
Lulu Qu obtained her Ph.D. degree from East China University of Science and Technology in 2014. Currently, she is an associate professor of Jiangsu Normal University.
Her research interests mainly include the preparation, characterization, and applications of biosensors.
Xiangmin Miao obtained her Ph.D. degree in Analytical Chemistry from Sun Yat-Sen
University, China, in 2012. Now, she is working in Jiangsu Normal University and
the current fields of interest were mainly biosensors, optical and electrochemical
nano-biological analysis and immunoassay.
Qiumei Feng obtained her Ph.D. degree in Analytical Chemistry from Nanjing University in 2016. Currently, she is an associate professor of Jiangsu Normal University.
Her research interests mainly include electroanalytical chemistry, biosensors, and
the synthesis and applications of nanomaterials.
Please cite this article in press as: P. Wang, et al., Construction of a paper-based electrochemical biosensing platform for rapid and
accurate detection of adenosine triphosphate (ATP), Sens. Actuators B: Chem. (2017), https://doi.org/10.1016/j.snb.2017.10.024
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