Nanotechnology and nanomaterial-based no-wash electrochemical biosensors: from design to application

Yong Zhang *ab and Xiaoyuan Chen *b
aKey Laboratory of Interfacial Reaction & Sensing Analysis in Universities of Shandong, School of Chemistry and Chemical Engineering, University of Jinan, Jinan 250022, PR China. E-mail: yongzhang7805@126.com
bLaboratory of Molecular Imaging and Nanomedicine (LOMIN), National Institute of Biomedical Imaging and Bioengineering (NIBIB), National Institutes of Health (NIH), Bethesda, Maryland 20892, USA. E-mail: shawn.chen@nih.gov

Received 6th July 2019 , Accepted 30th August 2019

First published on 3rd September 2019


Nanotechnology and nanomaterial based electrochemical biosensors (ECBs) have achieved great development in many fields, such as clinical diagnosis, food analysis, and environmental monitoring. Nowadays, the single-handed pursuit of sensitivity and accuracy cannot meet the demands of detection in many in situ and point-of-care (POC) circumstances. More and more attention has been focused on simplifying the operation procedure and reducing detection time, and thus no-wash assay has become one of the most effective ways for the continuous development of ECBs. However, there are many challenges to realize no-wash detection in the real analysis, such as redox interferences, multiple impurities, non-conducting protein macromolecules, etc. Furthermore, the complex detection circumstance in different application fields makes the realization of no-wash ECBs more complicated and difficult. Thanks to the updated nanotechnology and nanomaterials, in-depth analysis of the obstacles in the detection process and various methods for fabricating no-wash ECBs, most issues have been largely resolved. In this review, we have systematically analyzed the nanomaterial based design strategy of the state-of-the-art no-wash ECBs in the past few years. Following that, we summarized the challenges in the detection process of no-wash ECBs and their applications in different fields. Finally, based on the summary and analysis in this review, we also evaluated and discussed future prospects from the design to the application of ECBs.


image file: c9nr05696c-p1.tif

Yong Zhang

Yong Zhang received his BSc degree in applied chemistry from the University of Jinan, his MSc degree in applied chemistry from Nanjing University of Technology and PhD in school of materials science and engineering from Beijing Institute of Technology. He was appointed as an Associate Professor (2016) at the University of Jinan. He is now working as a visiting postdoctoral scholar of the National Institute of Health (USA) under the supervision of Prof. Xiaoyuan (Shawn) Chen. His research interests include preparing functional nanomaterials and developing new biosensors and bioelectronic devices.

image file: c9nr05696c-p2.tif

Xiaoyuan Chen

Xiaoyuan Chen received his Ph.D. degree in chemistry from the University of Idaho in 1999. He joined the University of Southern California as an Assistant Professor of Radiology in 2002. He then moved to Stanford University in 2004 and was promoted to Associate Professor in 2008. In the summer of 2009, he joined the Intramural Research Program of the NIBIB as a Senior Investigator and Chief of the LOMIN. He is interested in developing molecular imaging modalities for a better understanding of early diagnosis of disease, monitoring therapy response, and guiding drug discovery/development.


1. Introduction

Biosensors are a type of chemical sensor that is generally defined as analytical devices1–3 and have made great progress over the last three decades in various application fields that always involve all aspects of people's life safety and physical health, such as clinical diagnosis, food analysis, environmental monitoring, etc.4–6 Typically, there are two main systems that make a biosensor: a recognition system and a physicochemical transducer system,3,7 which also determine the analytical performance of the biosensor. The recognition system, which contains a biologically sensitive material, such as an enzyme, multi-enzyme system, antibody, organelle, bacterial or other cells,1 can specifically bind to or recognize the analyte or can stimulate a specific biological event with the introduction of the analyte.7 The physicochemical transducer system comprises electrodes, a signal converting and amplifying device, and a computer.3,8 During the process of detection, the physicochemical transducer system can detect signal changes within the recognition system on the interface of the electrodes, then convert and amplify the signals to electric signals, and finally provide a visualized spectrogram via computer software.9–11 As for the electrochemical biosensors (ECBs), the signals that change because of the alternation of the recognition system can be detected through one of the common electrical parameters, such as electric quantity, electric current, potential, or resistance.12

The rapid growth of nanoscience and nanotechnology provides great opportunities for the explosive development of ECBs that are constantly in pursuit for high sensitivity, high specificity and a low limit of detection (LOD).13–15 At present, the achievements in the sensitivity, selectivity, accuracy or LOD of ECBs have already been preparing to make the popular reality or commercial application of ECBs start to come true.6,7 However, people have been more and more aware that the single-handed pursuit of sensitivity and accuracy cannot meet the demands of the detection in many in situ or point-of-care testing (POCT) circumstances,16–18 especially in the fields of clinical diagnosis,19 food analysis,20 and environmental monitoring.21 More and more attention has been focused on simplifying the operation procedure and reducing detection time, and thus no-wash assay has become one of the most effective ways for the continuous development of ECBs.22–24

A no-wash biosensor assay refers to the detection performed just by simply mixing the sample solution and signal generating probes together, and then directly detecting signals generated from the mixture.5 Compared with conventional ECBs, no-wash ECBs have many advantages, such as simpler operation, faster and more convenient detection, etc., which make them more suitable for application in the in situ and POCT circumstances.25–28 In recent years, due to these merits, no-wash ECBs have been attracting more and more research interest and more and more studies have been carried out on them. With the development and deep study of research, many challenges have emerged for realizing no-wash detection in the real analysis, such as redox interferences,22 multiple impurities,29 non-conducting protein macromolecules,30 and so on. Furthermore, the complex detection circumstance in different application fields13,31–33 also makes the no-wash ECBs hard to achieve.

With the aid of interdisciplinary integration and the updated nanotechnology and novel functional nanomaterials,34–36 the in-depth analysis of the obstacles in the detection process and various methods for fabricating no-wash ECBs, the abovementioned issues have been addressed one by one. In this review, we have summarized the nanomaterial-based design strategies of the state-of-the-art no-wash ECBs in recent years. To be more specific, we will emphasize the functional impact and the role played by nanomaterials in the fabrication of no-wash ECBs instead of a specific type of nanomaterial, on which several review papers have been published more recently.7,13,37–40 Following that, we illustrated representative no-wash ECBs in different application fields. Finally, we also evaluated and discussed the challenges and future prospects from the design to the application of ECBs.

2. The design strategy of no-wash ECBs

Compared to liquid chromatography tandem mass spectrometry (LC-MS/MS), enzyme-linked immune sorbent assay (ELISA) and LC, the no-wash ECBs can realize portable, fast, accurate and sensitive detection with a simple operating process regardless of the special requirements of sample color and detection circumstance.41–44 These inherent advantages of electrochemical biosensors should be attributed to their test methods and detection mechanisms.45 Furthermore, by the combination of nanomaterials and nanotechnology, different types of ECBs make it reliable and possible for the no-wash electrochemical analysis to obtain high selectivity and specificity. In this review, we classified the recently published no-wash ECBs (or those have great potential to achieve no-wash detection) into four different types: signal change, ratiometric, direct reaction and others.

2.1. Signal change

The strategy of signal change is the most studied and easiest to realize in the ECBs’ design. Generally, there are three main types, i.e. signal-on,46 signal-off,47 and signal enhancement.48 For the no-wash ECBs, no matter what type of the strategy of signal change is, the key issue is to improve the signal-to-noise ratio (SNR) as much as possible. Therefore, among these three types, the signal-on strategy has the best sensitivity and greatest potential to fabricate no-wash ECBs under the same conditions, such as similar electrode materials, same method and same target analytes, as its design is based on the change of target induced signal change from zero to what to be read out. Nevertheless, the development of nanotechnology and the prepared functional nanomaterials also show the possibility of signal-off and signal enhancement strategies for fabricating no-wash ECBs.
2.1.1. Signal-on. The “signal-on” mode means that after the addition of the target analyte the signal response emerges due to the analyte triggered or induced specific reaction, such as immune interaction,49 DNA chain folding or cutting off,31 etc. Due to the signal rise from zero to a specific extent compared with the background signal in the absence of the target, this mode in theory can achieve a much improved signal response and obtain an increase in signal without limit.50

In principle, the nanomaterials used for the modification of the electrode or acting as a label have the capability of electrochemical redox activity themselves or trigging an electrochemical redox reaction. For example, Gu et al. developed a “signal-on” electrochemical biosensor based on the target-induced displacement reaction for detecting tumour cells.51 In that work, mesoporous silica nanoparticles (MSN) that were positively charged (PMSN) were used to entrap the electroactive probe (i.e. [Fe(CN)6]3−) into their pores. Then, the aptamer of the tumour cell with negative charge as a bio-gate was adsorbed on the surface of PMSN via the electrostatic interaction. As the electroactive probe was encapsulated in the pores of PMSN, only a small electrochemical signal was observed in the absence of the target tumour cell. Once the target tumour cells were added, they would be recognized and captured by the aptamer. As the adhesion between the bioconjugate and Apt-cell complex was decreased, the bio-gate was opened and the electroactive probes were released. Subsequently, the electrochemical signals emerged and significantly increased; thus the “signal-on” electrochemical biosensor was realized. Since such a “signal-on” strategy could avoid the drawbacks of “signal-off” strategies, the obtained LOD for tumor cells was 13 cells per mL. This assay was also extended for detecting microRNA (38 aM) and Hg2+ (0.47 pM).

It is well known that metal–organic framework (MOF) nanomaterials have great potential in the application of ECBs due to their large surface area for immobilization of biomolecules and electrochemical catalytic properties for acting as a redox label.52–55 However, still many issues remain, such as unsteadiness in aqueous environments and poor electron transfer capability. Therefore, many other nanomaterials, which have advantageous performances in one or two and more aspects, such as electrochemical activity, electrochemical catalytic properties, electron transfer capacity, and so on, and can act as the signal-on label or platform, have been prepared and utilized.56–63

In addition to the use of nanomaterials as the redox mediator, some chemical compounds containing redox active centers are also common for preparing ECBs, such as hemin, thionine (Thi), potassium ferricyanide, etc. Among them, hemin, known as a porphyrin, is one of the most used electron transfer mediators in various ECBs.64–67 It consists of an iron(III) ion, which is embedded in the center of a heterocyclic ring that contains four cyclically linked pyrrole molecules. Notably, unlike the other used redox mediators, the binding effect between hemin and oligonucleotide is noncovalent, and thus a hemin/G-quadruplex complex as a whole can serve as an electroactive molecule.66 Therefore, it provides a great opportunity for developing ECBs, which is based on proximity hybridization triggered hemin/G-quadruplex formation,50 and thus can design the signal-on strategy for no-wash ECBs.

Recently, Liu's group developed an ECB for the detection of lead ions (Pb2+) in tap water and pool water.64 As shown in Fig. 1, they designed and prepared three-dimensional DNA tetrahedra acting as a DNAzyme probe via adding a pendant substrate strand on a DNA tetrahedral nanostructure, and the substrate strand contains a G-rich fragment. In the process of detection, the conformational substrate strand in the presence of Pb2+ and then release the G-rich part. The hemin would subsequently combine with the release part and a G-quadruplex/hemin structure would be formed. This G-quadruplex/hemin structure provided an ultrasensitive electrochemical catalysis signal and thus the analysis by switch was displayed in two steps: DNAzyme would cleave the means of the signal-on no-wash ECB. Notably, the LOD of this ECB for the detection of Pb2+ is 0.008 nM, which is 6000 times and 9000 times lower than the minimum safety value of IARC (10 μg L−1 or 48.26 nM) and EPA (15 μg L−1 or 72 nM).


image file: c9nr05696c-f1.tif
Fig. 1 (A) Illustration of the label-free Pb2+ electrochemical biosensor based on the Pb-DNAzyme triggered G-quadruplex/hemin on a DNA tetrahedron. DG4 represents the side length of the hemin/G-quadruplex structure, and the DTDP represents the dimension of TSP. (B) CV results of Pb2+ analysis with the different concentrations. Reprinted with permission from ref. 64. Copyright 2018 Elsevier B.V.
2.1.2. Signal-off. In the “signal-off” strategy, the specific target binding would limit electron transfer or collisions between the sensing surface and the redox label thereby obtaining a decreased response current. Nevertheless, in the fabrication process of ECBs, the current may be further reduced because of the increased resistance, which may be caused by adsorbing many species (e.g. bacteria, virus, proteins, cancerous molecules, food pathogens, nonconductive or semiconductor materials, etc.) onto the sensing surface. Furthermore, for the no-wash ECBs, the nonspecific binding of molecules (e.g. antibodies, DNA, and biotins, etc.) with foreign molecules other than the target analytes is also an important issue in the fabrication of biosensors. In addition, the “signal-off” strategy might obtain false-positive results because of the coexistence of environmental stimulus.50 Therefore, the signal-off mechanism limits the gain of the sensor and delays the commercialization to some extent.

However, the incorporation of advanced nanomaterials into ECB development has further improved their signal-to-noise ratio and thus LOD, which provided great potential to achieve no-wash measurement via intelligent design and advanced nanotechnology. As shown in Fig. 2, Dutta et al. presented a signal-off strategy based wash-free and label-free immunosensor for the detection of Plasmodium falciparum histidine-rich protein 2 (PfHRP2), an important malaria protein biomarker, in whole blood.26 They used methylene blue (MB) as the medium substrate, Ru(NH3)63+ as an electron mediator, and an affinity-based sensing system as the protein quantifying portion where surface attachment of the target antigen effectively forms an insulator layer. In the presence of the target PfHRP2, the layer would impede electron transfer and thus distinctly reduce the electrochemical current. By applying this no-wash ECB, PfHRP2 could be measured from 0.1 to 100 μg mL−1 in whole blood samples without any steps of sample processing, modifying, labelling or washing.


image file: c9nr05696c-f2.tif
Fig. 2 Exploded view of the assembled immunosensor highlighting the major components. (A) Blood sample; (B) PET film with an inlet; (C) Vivid Plasma Separation membrane containing ascorbic oxidase; (D) cellulose membrane containing anti-PfHRP2 secondary antibody and methylene blue; (E) cellulose membrane containing Ru(NH3)63+; (F) Au electrochemical sensor (WE: working electrode, CE: counter electrode, and RE: reference electrode). Reprinted with permission from ref. 26. Copyright 2018 Nature Publishing Group.

Moreover, to overcome the instinct deficiency of the signal-off strategy, combining with the signal-on strategy has been studied widely, and was discussed in the section of ratiometric assay in detail.

2.1.3. Signal enhancement. As stated above, the amplification of electrochemical signals for the development of the no-wash ECBs must display extremely sensitive contrast to the background signals in the strategies of both signal-on and signal-off. Therefore, the strategy of signal loop amplification might have great potential for preparing no-wash ECBs. In the traditional DNA based ECBs, polymerase chain reaction (PCR) is one of the most common methods of signal loop amplification, and is used for the amplification of nucleic acid.68 However, the requirement of precise operation and cyclic temperature control limits its application in developing no-wash ECBs. Recombinase polymerase amplification (RPA) that is based on isothermal amplification is newly developed and suitable for the signal amplification for the fabrication of no-wash ECBs.25 The distinct advantages of RPA are that it does not need to control temperature precisely and does not require an initial denaturation step at an elevated temperature.69 To remove the washing or purification steps, Yang's group used a zinc finger protein, Zif268, to prepare no-wash ECBs based on the proximity-dependent electron mediation of ferrocenemethanol between the ITO electrode and the label of a glucose-oxidase (GOx), which can differentiate the specifically electrode-unbound and -bound labels without washing steps.25 The LOD for the detection of DNA of Piscirickettsia salmonis by this no-wash ECB was about 300 copies in 13.2 μL.

Notably, a method called electrochemical-enzymatic (EN) or electrochemical-chemical (EC) redox cycling, which means that a redox-active species as the proximity-dependent electron mediation allows faster electron transfer than an unbound one between an EN or EC label and an electrode, has been widely reported.22,70 In this redox cycling, there is a redox label for EN or EC redox cycling and an electron mediation for electron transfer, such as oxygen-insensitive diaphorase (DI) and Os(bpy)2Cl2,22 glycerol-3-phosphate dehydrogenase (GPDH) and Ru(NH3)63+,70 4-nitroso-1-naphthol and H3N–BH3,71 tris(2-carboxyethyl)-phosphine (TCEP) hydrochloride, p-aminophenol,72 etc. The difference between EN and EC redox cycling is that the EC redox cycling process always involves catalytic nanomaterials, such as noble metal NPs, reduced graphene, transition metal nitride, etc.

In addition to the above signal loop amplification methods based on the polymerase reaction and EN or EC redox reaction, Park's group used a personal glucose meter (PGM) to develop a no-wash and label-free assay based on the enzymatic reaction for the detection of adenosine 5′-triphosphate (ATP) in human serum and bovine urine.27 There are two steps of reaction in the process of their proposed assay. First, the glucose in the blood is converted to glucose 6-phosphate by the catalysis of hexokinase in the presence of target ATP. As a result, the amount of ATP in the blood is proportional to the decrease of the amount of glucose. Then, the enzymatic reaction of hexokinase will produce adenosine 5′-diphosphate (ADP), which will be recovered to ATP, catalyzed by pyruvate kinase enzyme. The reproduced ATP is again provided to catalyze multiple loops of enzymatic reactions, thus leading to a great signal amplification.

2.2. Ratiometric assay

Different from the signal change method, which is only dependent on one signal, the ratiometric assay is designed from a built-in self-calibration of signal intensity by means of recording at least two target-induced signal fluctuations.73–75 Therefore, the ratiometric electrochemical assay may obtain greater accuracy, reproducibility, and sensitivity than the classic electrochemical assay.76,77 Generally, there are three strategies to achieve ratiometric electrochemical detection. One strategy is to introduce the second signal as a reference that is target-insensitive for signal correction. The second one is to apply two target-responsive reversible signal changes that enable the ratiometry. The third strategy is to use two target-responsive labels that they can respond in concert to the environmental changes that cause drift.
2.2.1. Reference signal correction. The principle of the strategy of reference signal correction is that the signal correction relies on the utilization of two different redox reporters conjugated to one aptamer probe for electrochemical aptamer-based (E-AB) biosensors78 or electrochemically respond to the same target molecule for electrochemical immunosensors.79,80 In other words, in these response signals, one acts as a reference factor to calibrate the others. Without worrying about the artificial factors and electrode modification layer, etc.,81 this strategy can greatly reduce or possibly overcome the measurement differences of the detection environment, and thus as far as possible to provide a test result with better accuracy, reproducibility and reliability.74,82 Therefore, an appropriate reference label, which can generate not only a single and stable response signal but also a distinct response signal from the counter signals, is very essential. Until now, several redox mediators have been used as redox reporters to provide a reference signal, such as Methylene Blue (MB, redox current peak at ∼−0.28 V),80,83,84 Thi (∼−0.23 V),85 Nile Blue (NB, ∼−0.325 V),86 Ferrocene (Fc, ∼+0.47 V),87 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonate) (ABTS, ∼+0.5 V),88 etc. Since these redox substances have a stable signal output and can be obtained easily, the only thing for designing the ratiometric strategy is to explore and find another redox substance out. The emerging advanced nanomaterials provide us more choices for employing redox reporters with perfect performance either as the reference label or as the counter label, such as noble metal nanomaterials,89 MOFs,90,91 carbon nanomaterials,92 quantum dots (QDs),61,93 porphyrin compounds,52 transition metal oxides,94 their compositions,80 etc.

For example, to continuously measure multiple analytes in whole blood, Zhang et al. prepared dual-function MXene with a morphology of 2D nanosheets,95 in which the MXene is exfoliated from their ternary carbide and nitride ceramics and has been studied in many fields.96–99 In that work, as shown in Fig. 3, MXene was used to modify the electrode to increase the electrode surface area and thus improve the catalytic activity. MB loaded on MXene acted as a reference-labelling molecule to adjust the electrochemical background signal. Urease immobilized on MXene was able to catalyse urea with the aid of glutaraldehyde to generate specific and distinct signals from MB towards the selective and sensitive detection of urea. Due to the stable reference signal and the sensitive signal change according to the concentration of the target analyte, the no-wash ECB achieved accurate and stable measurement of urea in whole blood with a linear range from 0 to 3 × 10−3 M.


image file: c9nr05696c-f3.tif
Fig. 3 (a) Schematic illustration of the preparation process of the MXene-based microfluidic chip. (b) Schematic illustration of detection of uric acid at the MXene modified electrode. (c) The square wave voltammetry plots toward different concentrations of uric acid. Reprinted with permission from ref. 95. Copyright 2019 WILEY-VCH Verlag GmbH & Co. KGaA.

Moreover, unlike the traditional ratiometric electrochemical measurement via only one electrochemical method, e.g. differential pulse voltammetry (DPV), the ratiometric ECBs can be designed by means of two different signal transducers. In other words, such assay is developed by combining with two different electroanalytical technologies.101 Recently, Lin et al. have integrated electrochemical technology with electrochemiluminescence (ECL) technology to develop a ratiometric biosensor for the detection of Golgi protein 73 (GP73), a hepatocellular carcinoma (HCC) biomarker, in human serum.102 In that work, they utilized MB as the electrochemical redox reporter and tris-(4,4′-dicarboxylicacid-2,2′-bipyridyl) ruthenium(II) dichloride (Ru(dcbpy)3Cl2) as the ECL signal reporter. To be more specific, the MB, which was carried by the composed double-stranded DNA (dsDNA), provides a stable electrochemical internal reference signal on the electrode surface to correct the potential interferences. Simultaneously, Ru(dcbpy)3Cl2, which was modified on the electrode through the formation of the classical sandwich complex of the immunosensor (i.e. antibody–antigen–antibody), provides increased ECL response signals with the concentration increment of GP73. During the measurement, the ratio value of the ECL signal of Ru(dcbpy)3Cl2 and the current of MB showed a linear relationship with GP73 in the concentration range of 15 pg mL−1–0.7 ng mL−1 and the LOD was 15 pg mL−1.

2.2.2. Two-signal ratiometry. In this mode, the two redox reporters will simultaneously generate changing signals in accordance with the amount of the target analyte, such as both of signal-on reporters85 and signal-on reporter combined with a signal-off reporter.80,103

For example, Zhu et al. proposed a ratiometric E-AB biosensor for highly sensitive detection of thrombin (TB).100 As shown in Fig. 4, firstly, they prepared a double-stranded DNA via hybridizing the thrombin aptamer (TBA) and DNA walkers (DWs). Following that, they immobilized both MB labelled DNA (MB-DNA) and DWs/TBA double-stranded DNA onto the Au NP modified glass carbon (GC) electrode. After adding ferrocene-labelled DNA (Fc-DNA), a MB-DNA/Fc-DNA duplex would form because of the hybridization of the MB-DNA with the Fc-DNA. During the detection, in the presence of the target TB, DWs could be released because of the specific interaction between TB and TBA, and the released DWs subsequently hybridized with part of Fc-DNA to form a duplex with a blunt end at the 30-termini of Fc-DNA. Then, the Exo III cleavage process was accompanied by throwing away Fc from the electrode and triggering the release of DWs. Finally, the DWs continued to hybridize another Fc-DNA to perform a recycling process, and a hairpin structure in the presence of Mg2+ could be further formed by the remaining single-stranded MB-DNA. Therefore, a suppression of the Fc signal (IFc) and an enhancement of the MB signal (IMB) were achieved. Based on the ratio of IMB/IFc, this E-AB biosensor showed superior analytical performance for the detection of TB with a LOD of 56 fM.


image file: c9nr05696c-f4.tif
Fig. 4 Schematic illustration of the E-AB biosensor for the detection of TB. Reprinted with permission from ref. 100. Copyright 2018 The Royal Society of Chemistry.
2.2.3. Signal drift correction. The strategy of signal drift correction has the same principle as the strategy of reference signal correction except that the two different redox reporters have sufficiently similar physical and stable properties under the same analysis environment so that they can respond in concert to the signal drift caused by the environmental changes.

Li et al. used anthraquinone (AQ) as a reference reporter and MB as a sensing reporter to fabricate a no-wash E-AB sensor based on dual-reporter drift correction.104 As shown in Fig. 5, they placed MB on the distal end of the aptamer and AQ on the nucleobase proximal to the electrode. Since the MB (i.e. the sensing reporter) is away from the electrode, it can approach the electrode to transfer electrons through target induced conformation (i.e. binding-defined) of the aptamer. In contrast, because the AQ (i.e. the reference reporter) is proximal to the electrode, its distance from the electrode is not highly dependent on the conformation of the aptamer. Therefore, while both the reference and sensing reporters respond to the drift caused by environmental effects, only the signal generated from the sensing reporter can respond to the target.


image file: c9nr05696c-f5.tif
Fig. 5 (A) Schematic of the E-AB sensor based on the dual-reporter drift correction. (B) Voltammograms of MB and AQ that respond to the target analyte. (C) Both redox reporters’ drifts are illustrated when the sensor is placed in flowing, undiluted whole blood. (D) The currents from the two reporters likewise drift in concert when the sensor is challenged with varying concentrations of its target in flowing whole blood. (E) After correction, the drift decreases significantly, recovering target concentration estimations (green curve) in close agreement with the actual concentrations of the drug spiked into the sample (red trace). Reprinted with permission from ref. 104. Copyright 2016 American Chemical Society.

In brief, although good analytical performance of ratiometric ECBs has obtained by means of advanced nanomaterials, not all the functional nanomaterials are appropriate for fabricating no-wash ECBs. To meet the requirements for no-wash detection, especially in the flow liquid environment, the nanomaterials used as redox reporters should possess three elemental properties, that is, good stability in a complex physiological or natural environment, excellent electrochemical redox activity instinct or to the other enzymes at the nearest potential to 0 V, and friendly biocompatibility for the incorporation with the biomolecule that can specifically recognize the target.

2.3. Direct reaction

Different from the above two kinds of ECB strategies, in which the signals come from functional nanomaterials or biological species with electrochemical activities other than the target analyte, the direct reaction mode means that the target analyte directly involves in the reaction and obtains an specific electrochemical signal. To propose and achieve no-wash detection, there are two forms in this strategy mode in general.
2.3.1. Signal generated from the direct reaction of the target. In this process of determination, the obtained signal is provided by the electrochemical redox reaction between the target analyte and the sensing electrode. Since there are always various redox species in the complicated sample environment, it is in general hard to acquire the exact intensity of the specific electrical signal of the target analyte.106 Therefore, similar to the above described ratiometric strategy, it is effective for no-wash detection by introducing a reference signal and testing the ratiometric intensities of dual electrical signals generated from the target and the reference species. Recently, a selective ratiometric ECB for accurate determination of Hg2+ in environmental water was developed by Ma et al.107 In that work, MB acted as the reference label, and Hg2+ and MB would generate redox peak currents at +0.59 V and −0.2 V, respectively. To be more specific, phenyl thiourea (PT) was used as a specific recognition element for adsorbing Hg2+, and (5′-MBAGGAGGAGGAGGGAGGAGGG-SH-3′ (HS-DNA-MB) was designed as an inner reference element. In the process of detection, the redox peak current at +0.59 V gradually increased with the increase of the concentration of Hg2+; meanwhile the redox peak current at −0.2 V remained almost constant. The peak current density ratio at these distinct potentials demonstrated a good linearity with the concentration of Hg2+ in a broad range from 1 nM to 1.3 μM and the LOD was down to 0.12 nM, which is much lower than the standard values required by the WHO and EPA. With the aid of MB as the reference label too, Yu et al. prepared a facile ratiometric electrochemical sensor for selective and sensitive determination of doxorubicin hydrochloride (DOX), a kind of chemotherapy medication used to treat cancer.106
2.3.2. Signal generated from the reaction induced by the target. In this process of determination, the target analyte will not generate electrochemical redox signals. The obtained signal comes from the other reaction triggered by the target than the target itself. It means that an extremely obvious electrochemical signal current vs. background signals emerges as the target analyte triggers a special reaction. In order to realize no-wash measurement, the generated current signal should be stable and have a great signal-to-noise ratio. Therefore, choosing an exactly special reaction and a signal generator with good performance are very important. For example, Li et al. reported a target-driven E-AB biosensor for the real-time and continuous detection of melamine in flowing milk.105 As shown in Fig. 6(A), they used MB modified and thiol-anchored DNA sequences to construct two polythymine segments, which were connected by a four-cytosine loop. This four-cytosine loop can vary from 4 to 12 bases. By tuning the length of polythymine segments, they could tune the extent of cooperativity. In the presence of melamine, a conformational change occurs when there is binding between melamine and bases. Such conformational change alters the ability of the redox reporter to approach the electrode, and produces a target-dependent change in current when the sensor is interrogated via square wave voltammetry. When challenged with 500 μM melamine undiluted whole milk, a significant signal change in Fig. 6(B) and rapid binding event in Fig. 6(C) could be observed. Thus, the no-wash and real-time measurement of melamine ECBs was successively developed.
image file: c9nr05696c-f6.tif
Fig. 6 (A) Schematic of the E-AB sensor supporting no-wash, real-time and continuous detection of melamine in whole milk. (B) Square-wave voltammetry (SWV) scan diagram and (C) binding rate diagram in the presence and absence of 500 μM melamine in undiluted whole milk. Reprinted with permission from ref. 105. Copyright 2018 American Chemical Society.

2.4. Others

In addition to the above strategy, there are still some other less commonly used methods designed for preparing no-wash ECBs. Among them, the magnetic beads (MBs) maybe the most used nanomaterials46,108 and separation of MBs that label the target analyte from the sample solution may be the most popular nanotechnology.109 The reason is that the MBs can be easily collected by means of a magnet, and thus can be used as a carrier for the pre-concentration or specific capture of different molecules in sensing systems to improve the sensitivity and the resistibility against matrix interference of the method.46

Direct electron transfer between a redox reporter and the electrode without the participation of electron mediators is also an option for designing the no-wash ECB strategy. In this strategy, an affinity biosensor based on direct electron transfer generally requires a finely smoothed gold electrode to support efficient target binding, and a short working distance (<1–2 nm) between a redox reporter and the electrode.30,110,111 However, an anionic π-conjugated polyelectrolyte (CPE) label has been reported recently to achieve a long-range electron transfer between a diffusing redox-active species and an electrode.30 In that work, after sandwich-type target-specific binding, the long CPE label (∼18 nm for 10 kDa) can approach the electrode within the working distance, and fast CPE-mediated oxidation of ammonia borane along the entire CPE backbone affords high signal amplification. Thanks to the high signal amplification, the proposed ECBs succeeded to detect the target DNA with a LOD of 10 pM without a washing step.

3. Applications and outlooks of no-wash ECBs

3.1. The issues that need to be faced before applications

Although various no-wash ECBs have been fabricated in recent years, there are still some limitations that impede the real-world application of no-wash ECBs, for example, signal drifting due to the change in environmental conditions, electrode surface passivation and contamination after exposure to samples, decreased stability of the electrode after introducing extra sensing agents, reduction of electronic transmission capacity after modification, etc.

For example, in the field of clinical diagnosis, blood is the most important test sample. Blood is the body fluid that transports essential substances such as oxygen, nourishment and hormones throughout the body. Blood also delivers disease fighting substances and metabolic waste products from cells.112–115 For the clinical diagnosis, blood is simple to process and readily available.116 In routine clinical analysis, blood will be pre-treated to obtain plasma and serum depending on the different analytical applications and detection types.117

As for the fabrication of ECBs, there are two main types of interferences in the real sample measurement. One is common to all kinds of ECBs, which is a similar substance to the target analyte in one or more aspects, such as chemical properties, morphology, biological nature and biological activity. The other interference in the real sample measurement is redox-active species. This type of interference makes the no-wash electrochemical detection more complicated. Recently, Yang et al. classified these redox-active interfering species into several types.22 As shown in Fig. 7, one type of interference is generated from the electrochemical oxidation of interfering species (ISs) at a sensing electrode (schemes (i), (ii), and (iii) in Fig. 7). The second type of interference comes from slow electrochemical oxidation of an electron mediator (M) because of partial electrode passivation or strong interaction with ISs (schemes (iv) and (v) in Fig. 7). The third type of interference is generated from redox reactions between ISs (or O2) and a redox enzyme label (scheme (vi) in Fig. 7). The last type of interference is generated from redox reactions between ISs (or O2) and an electron mediator (scheme (vii) in Fig. 7). Generally speaking, to avoid nonspecific adsorption of biological substances other than the target analyte, one can use a blocking reagent, such as bovine serum albumin (BSA)3,7,118 and oxidized glutathione (GSSG),119 to block those nonspecific active sites on the surface of the ECBs. However, the blocking agent alone is not enough to achieve no-wash detection. For example, when an immunosensor with an enzyme-labelled antibody is bound to a target immobilized on the sensing surface by immune interaction, the sensing surface may non-specifically adsorb this labelled antibody, which results in a high zero signal and low change of the target-induced signal.22 Therefore, minimizing the nonspecific adsorption of the antibody that is labelled for generating signals is also very important for fabricating no-wash ECBs.


image file: c9nr05696c-f7.tif
Fig. 7 Schematic of possible interfering reactions. (i), (ii), and (iii) are the interferences generated from the electrochemical oxidation of interfering species (ISs) at a sensing electrode. (iv) and (v) are the interferences generated from slow electrochemical oxidation of an electron mediator (M) because of partial electrode passivation or strong interaction with ISs. (vi) is the interference generated from redox reactions between ISs (or O2) and a redox enzyme label. (vii) is the interference generated from redox reactions between ISs (or O2) and an electron mediator. Reprinted with permission from ref. 22. Copyright 2018 American Chemical Society.

3.2. The applications of no-wash ECBs

Nowadays, people have been more and more concerned about disease diagnosis, food safety and environmental pollution. For better protection of human health, numerous published research studies were involved in the rapid and accurate detection of various targets related to people's life and health, such as biomarkers, pathogens, bacteria, heavy metals, food and environmental pollutants, and so on. Thanks to their instinct merits as stated above, no-wash ECBs have been attracting increased attention in many fields, such as clinical diagnosis, food analysis and environmental monitoring.
3.2.1. The applications in clinical diagnosis. As an invaluable tool for the specific and sensitive detection of manifold biochemical species in clinical diagnosis, no-wash ECBs have been widely used to monitor blood glucose over the past decades. Until now, to meet the requirements of the measurements in the POCT or in situ circumstance, many research groups have developed a variety of no-wash ECBs for diagnosis of life-threatening diseases and for the detection of many substances related to people's physical health, such as biomarkers, bacteria, virus, proteins, cancerous molecules, DNA and so on.120–122 Since several research studies about the detection of biomarkers, DNA and other biomolecules have been illustrated in the previous sections, some other target analytes that emerge in common clinical diagnosis will be discussed in this section.

Depending on the change of the probe coverage on the sensing electrode, Wu et al. fabricated a no-wash ECB for real time and sensitive detection of cis-diamminedichloroplatinum-(II) (cisplatin), which is generally used as a front-line treatment against neoplasia, in particular ovarian and testicular cancers, and neck and head cancers in clinical settings.123 As shown in Fig. 8, they used methylene blue (MB)-modified oligo-adenine (A)-guanine (G) DNA as the probe. According to the probe coverage intensity on the electrode and depending on preferential formation of intrastrand cisplatin-AG, the ECB generated a change effect of signal-off or signal-on. Due to the distinct signal changes achieved, this ECB was realized to detect cisplatin in realistically complex samples, saliva and urine. The linear dynamic range was from 0.5 to 5 μM with a LOD of 500 nM.


image file: c9nr05696c-f8.tif
Fig. 8 Schematic illustrations of the high probe coverage signal-off (A) and low probe coverage signal-on (B) electrochemical cisplatin Sensors. Reprinted with permission from ref. 123. Copyright 2017 American Chemical Society.

Lately, Parate et al. fabricated a no-wash electrochemical sensor for the detection of cotinine in human saliva.124 Cotinine is a kind of metabolite of nicotine, which was used as a biomarker for the measurement of smoke exposure and tobacco use. To achieve no-wash detection of cotinine, they firstly modified the electrode with graphene and Pt NPs (Pt@graphene) and then electrodeposited the molecularly imprinted polymer (MIP), in which cotinine was the template molecule, onto the modified electrode. This modified electrode generated a 4-fold increase current intensity in ferricyanide solution via cyclic voltammetry (CV) scanning. As a result, this proposed ECB can specially and consequently detect cotinine in the sample of spiked saliva within a wide linear range of 1–100 nM and the LOD is 0.33 nM.

The intracellular redox homeostasis is able to regulate cell micro-environmental reactions and thus the cell behaviour and function.125 Recently, thanks to the high affinity between DNA and metal ions and the high affinity between GSH and Ag NPs, Liu et al. reported a ratiometric ECB based on the DNA hybridization chain reaction (HCR, an enzyme-free amplification method) by metallization mediated for determining Glutathione (GSH) for intracellular redox homeostasis monitoring.126 Firstly, a water-soluble porphyrin (TCPP) was modified onto the electrode to stabilize chemically converted graphene (CCG), which possessed high conductivity. Due to the abundant negatively charged carboxyl groups on CCG, NH2-modified hDNA could be linked and retained on the CCG and the unreacted NHS esters were passivated by ethanolamine. Following that, Ag NPs were deposited onto the conjugated hDNA, and thus DNA base pairing could be disturbed and DNA hybridization was further hindered. In the presence of GSH, the Ag NPs would be released since Ag NPs and GSH have a stronger affinity, which induced a remarkable current reduction via the reaction of Ag to AgCl (IAg). Meanwhile, the in situ HCR was triggered by the liberated hDNA to produce self-assembled long fragments of DNA. After the addition of MB, which could intercalate into dsDNA and generate a signal-on current (IMB), the HCR process was monitored and the GSH could be further measured by recording the current value ratio (IMB/IAg). Based on this ratiometric method, the LOD of the GSH ECB is 103 mM. In addition, normal cells and cancer cells could also be discriminated by means of this assay.

3.2.2. The applications in food analysis. Nowadays, a high degree of automation and continuous processing are gradually replacing many batch operations in the food industry.127,128 Accordingly, the demands for instruments more and more need to be suitable for automatic quality control in the whole process of the production line. Therefore, more concern on the in situ test or on-line monitoring rather than overemphasizing the LOD is different from clinic diagnosis. Until now, several research studies on no-wash electrochemical sensors or biosensors have been reported.

Xu et al. fabricated a dual-signal E-AB for the detection of Malathion in cauliflower and cabbage.129 Malathion is a kind of common organophosphorus insecticide, which may cause muscle dysfunction, Alzheimer's disease and thyroid cancer if people ingest foods with its residues. As shown in Fig. 9, they firstly prepared the poly-dopamine (PDA)-Au NP (PDA-Au NP) composition with good electrical conductivity and biocompatibility via electrodeposition onto the electrode directly. Afterwards, in the presence of Malathion, the specific aptamer to Malathion would be detached from the electrode and then a hairpin structure would be formed induced by the capture probe. Following that, exonuclease I (Exo I) was added to motivate the autocatalytic target cycling reaction and obtained a dramatically increased electrochemical current. Based on the signal-off of thionine (Tn) and the signal-on of ferrocene (Fc), the proposed E-AB provided a dual-signal current readout. Under the optimized conditions, this E-AB for the determination of Malathion exhibited a linear range of 0.5–600 ng L−1 with a LOD of 0.5 ng L−1.


image file: c9nr05696c-f9.tif
Fig. 9 Schematic illustration of the dual-signal aptamer sensor based on PDA-Au NPs and Exo I signal amplification for the detection of malathion. Reprinted with permission from ref. 129. Copyright 2019 Elsevier B.V.

For the detection of another kind of organophosphorus insecticide, methyl parathion, Wang et al. proposed a no-wash ECB based on the direct and specific reaction between the target analyte and Burkholderia cepacia lipase (BCL) on the sensing electrode.130 For carrying BCL, Cu based MOF nanofibers were prepared at first.

After the BCL was loaded onto MOF nanofibers and further immobilized onto the electrode with the aid of chitosan, the ECB was obtained. In the process of detection, the ECB was directly inserted into the sample solution containing the target methyl parathion and monitored via the DPV method. The BCL would catalyse hydrolysis of methyl parathion and obtain a remarkable redox current signal at about −0.11 V. This ECB achieved direct detection of methyl parathion in vegetable samples without washing steps within the linear range of 0.1–38 μM and the LOD was 0.067 μM.

Based on a target-induced signal-shifting of the labelled probe, Li et al. developed a signal-on E-AB biosensor for the determination of kanamycin in milk, tap water and serum without washing steps.131 To design the signal-on strategy, three probes were used in their study. A thiolated assistant probe and a no-thiolated capture probe were all immobilized onto the sensing electrode. The third probe was an MB-labelled probe for providing signals and mixed with the sample solution. In the absence of kanamycin, the background signal was extremely small as the MB-labelled probes were far away from the electrode and provided low efficient electron transfer. Nevertheless, in the presence of kanamycin, since the stable complex of the no-thiolated capture probe/kanamycin complex was formed and thus the former short duplex was undone, the assistant probe was released. Simultaneously, the free MB-labelled probe was combined with another assistant probe. This combination made MB shift close to the electrode and provided great highly efficient electron transfer. Therefore, a significant signal related to the target concentration was generated and the signal-on strategy was achieved. This E-AB biosensor features a high sensitivity with a LOD of 3.3 pM for the detection of kanamycin.

3.2.3. The applications in environmental monitoring. Different from clinical diagnosis and food analysis, which are operated in general under mild environmental conditions, environmental monitoring always requires emergency in situ detection and continuous measurement under harsh and robust conditions. Therefore, the no-wash electrochemical sensors or biosensors need to be more robust and endurance for the fast, in situ and low-cost detection than in the other application fields. During the past several years, not too many research studies have been published involving the preparation and application of no-wash electrochemical sensors or biosensors.

As for the demands of the harsh and robust detection environment, the technology of molecular imprinting (MI) offers a versatile, sturdy and effective method to improve the electrochemical sensor's surface enrichment performance.132–135 Therefore, a MIP based electrochemical sensor may have great potential for fabricating no-wash sensors to meet the requirements of environmental monitoring. Pei et al. designed a Bisphenol A (BPA) electrochemical sensor based on the combination of TiO2 single crystals (SCs) and MIP.137 BPA is a kind of persistent organic pollutant widely present in the environment and has severe health risks to humans. As it has an electro-active phenolic group, BPA is very suitable for determination via an electrochemical method. In that study, since the special shape- and facet-engineering structure of TiO2 SCs enhanced the reduction of their catalytic activity and electric conductivity and thus providing minimum background signals, a remarkable redox current signal and a great signal-to-noise ratio would be obtained by adsorption of BPA with the aid of inorganic-framework MI technology. This electrochemical sensor can achieve the linear detection of BPA within the range of 0.01–20.00 μM with a LOD of 3.0 nM.

Microfluidic devices have been attracting more and more attention in the field of analytical chemistry because of their inherent advantages, such as high speed of analysis, low consumption of samples and reagents, portability etc.138 To meet the requirement of environmental monitoring, as shown in Fig. 10, Caetano et al. fabricated a commercial textile thread based microfluidic device with the aid of the electrochemical method for detecting phenol in tap water, and the LOD was 2.94 nM.136 The paper-based platform is another noticeable technique, which has great potential to meet the requirement of the POCT.139–142 More recently, Moscone et al. reported a 3D origami paper-based platform for designing different enzyme-inhibition electrochemical biosensors. By means of these devices, they realized no-wash detecting several classes of pesticides only via folding and unfolding the filter paper-based structure, such as paraoxon, 2,4-dichlorophenoxyacetic acid, and atrazine. They evaluated the accuracy of this paper-based electrochemical biosensor in the sample of river water and obtained satisfactory results.143


image file: c9nr05696c-f10.tif
Fig. 10 Dimensions and constituents parts μ-TED (a) inlet reservoir, (b) injection zone, (c) SPE (electrochemical detection zone), (d) hydrophilic textile thread, (e) adhesive tape and (f) outlet reservoir. To demonstrate the solution flux on threads, a food colouring was employed. Reprinted with permission from ref. 136. Copyright 2018 Elsevier B.V.

Recently, Li et al. have reported a MOF based electrochemical sensor for the detection of catechol (CT) and hydroquinone (HQ), which are two common environmental coexisting dihydroxybenzene isomers (DBIs) of phenolic compounds, in tap water.144 They prepared a composite of the copper-based MOF and graphene (Cu-MOF/GN) and used it to adsorb and catalyse HQ and CT. During the DPV scanning, CT and HQ exhibited a significant electrochemical redox current peak separately and the LOD for the detection of CT and HQ was 330 nM and 590 nM, respectively. However, as the redox peak potentials of CT and HQ were both within a narrow range of 0.0–0.15 V, this senor succeeded in determining CT and HQ in the same platform instead of simultaneous detection.

3.3. The outlook of no-wash ECBs

The distinct merits of ECBs have been keeping them under focus in the fields of analytical chemistry from the past to the future. To push the ECBs into realization in commercial application, no-wash ECBs have shown great potential and achieved numerous advances in the past few decades. However, before the applications in the fields of clinical diagnosis, food analysis and environmental monitoring are realized, there are still a long way to go and some challenges that required to be addressed.

Firstly, the stable sensing platform with good reproducibility still needs to be improved. Not only for the test in the purified samples, but also for the measurement in the original complex samples, a stable substrate is very essential. Furthermore, for the no-wash detection, the phenomenon of signal drift and current correction always emerges, especially in the real-time and on-line sample analysis.

Secondly, the specific recognition mechanism still requires a deep study to further achieve excellent selectivity for accurately recognizing the target molecules in complex real samples. In particular, advanced nanomaterials and nanotechnologies still need to be explored for reducing the nonspecific adsorption on the sensing platform surfaces in a complicated measurement environment.

Finally, although various functional nanomaterials have been used to prepare the no-wash ECBs and great analytical performance of ECBs has been achieved, mass-scale controllable synthesis of nanomaterials for obtaining mass production of repeatable ECBs is still a matter of concern.

Based on the analysis and summary in this review, to improve the analysis performance, further investigation and study may focus on several aspects.

(i) Advanced nanomaterials with great and specific electrochemical catalytic properties need to be prepared to enhance the signal-to-noise ratio. Depending on this, novel ratiometric or signal-on ECBs could be designed to meet the requirement of no-wash detection in a physiological environment, especially in the fluid samples.

(ii) Novel specific recognition mechanism needs to be explored and studied deeply. So that, it will broaden the applications of no-wash ECBs for the detection of more and more species. In addition, applying a reliable and advanced recognition mechanism to improve the electrode modification process and reduce the nonspecific adsorption can obtain a high signal-to-noise ratio.

(iii) A more stable sensor substrate may be designed and prepared to obtain a reproducible and steady interfacial reaction and sensing analysis, such as nanoarray electrode, microfluidic chip and so on. Additionally, in the detection of multiple target analytes, which may involve multiple labelling signals in the assay, a stable sensor is the premise.

4. Conclusions

Over the past decades, the no-wash ECBs have been providing great potential for target analysis and measurement in various application fields, such as clinical diagnosis, food analysis, and environmental monitoring. Due to the fast development of nanotechnology and nanomaterials, there are many skilful and versatile detection strategies being designed and applied in the fabrication of no-wash ECBs, such as signal change, ratiometric and direct reaction, etc. Nevertheless, there are still some challenges to realize no-wash detection in the real analysis, such as redox interferences, multiple impurities, non-conducting protein macromolecules, etc. Furthermore, the complex detection circumstance in different application fields makes the realization of no-wash ECBs more complicated and difficult. In conclusion, although there is still a long way to explore, based on the great achievements and breakthroughs obtained previously in the preparation of no-wash ECBs, the mass production and popular application of no-wash ECBs in wide fields will be realized in the near future.

Conflicts of interest

The authors state that “There are no conflicts to declare”.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 21775053), the Shandong Provincial Natural Science Foundation (No. ZR2017MB027 and 2019GSF111023), and the intramural research program of the National Institute of Biomedical Imaging and Bioengineering (NIBIB), National Institutes of Health (NIH), and Yong Zhang is sponsored by the China Scholarship Council (CSC).

Notes and references

  1. C. R. Lowe, Trends Biotechnol., 1984, 2, 59–65 CrossRef CAS.
  2. D. R. Thévenot, K. Toth, R. A. Durst and G. S. Wilson, Biosens. Bioelectron., 2001, 16, 121–131 CrossRef.
  3. D. Grieshaber, R. MacKenzie, J. Voros and E. Reimhult, Sensors, 2008, 8, 1400–1458 CrossRef CAS PubMed.
  4. W. W. Zhao, J. J. Xu and H. Y. Chen, Chem. Soc. Rev., 2015, 44, 729–741 RSC.
  5. X. Huang, Y. Liu, B. Yung, Y. Xiong and X. Chen, ACS Nano, 2017, 11, 5238–5292 CrossRef CAS PubMed.
  6. T. Xiao, F. Wu, J. Hao, M. Zhang, P. Yu and L. Mao, Anal. Chem., 2017, 89, 300–313 CrossRef CAS PubMed.
  7. G. Maduraiveeran, M. Sasidharan and V. Ganesan, Biosens. Bioelectron., 2018, 103, 113–129 CrossRef CAS PubMed.
  8. S. Liu, H. Cao, X. Wang, W. Tu and Z. Dai, Nanoscale, 2018, 10, 16474–16478 RSC.
  9. Y. Zhang and Q. Wei, J. Electroanal. Chem., 2016, 781, 401–409 CrossRef CAS.
  10. Z. Liu, Y. Zhang, J. Feng, Q. Han and Q. Wei, Sens. Actuators, B, 2019, 287, 551–556 CrossRef CAS.
  11. C. Gao, Z. Guo, J. H. Liu and X. J. Huang, Nanoscale, 2012, 4, 1948–1963 RSC.
  12. C. S. Pundir, S. Jakhar and V. Narwal, Biosens. Bioelectron., 2019, 123, 36–50 CrossRef CAS PubMed.
  13. G. K. Mishra, V. Sharma and R. K. Mishra, Biosensors, 2018, 8, 28 CrossRef PubMed.
  14. R. Munter, K. Kristensen, D. Pedersbaek, J. B. Larsen, J. B. Simonsen and T. L. Andresen, Nanoscale, 2018, 10, 22720–22724 RSC.
  15. R. R. S. Packard, Y. Luo, P. Abiri, N. Jen, O. Aksoy, W. M. Suh, Y. C. Tai and T. K. Hsiai, Theranostics, 2017, 7, 2431–2442 CrossRef CAS PubMed.
  16. P. Kassal, M. D. Steinberg and I. M. Steinberg, Sens. Actuators, B, 2018, 266, 228–245 CrossRef CAS.
  17. J. Liu, Y. Wada, M. Katsura, H. Tozawa, N. Erwin, C. M. Kapron, G. Bao and J. Liu, Theranostics, 2018, 8, 6053–6069 CrossRef PubMed.
  18. H. Liu, X. Zhou, Q. Shen and D. Xing, Theranostics, 2018, 8, 2289–2299 CrossRef CAS PubMed.
  19. N. T. Darwish, S. D. Sekaran and S. M. Khor, Sens. Actuators, B, 2018, 255, 3316–3331 CrossRef CAS.
  20. N. F. D. Silva, J. Magalhaes, C. Freire and C. Delerue-Matos, Biosens. Bioelectron., 2018, 99, 667–682 CrossRef CAS PubMed.
  21. S. Uniyal and R. K. Sharma, Biosens. Bioelectron., 2018, 116, 37–50 CrossRef CAS PubMed.
  22. P. Nandhakumar, A. J. Haque, N. S. Lee, Y. H. Yoon and H. Yang, Anal. Chem., 2018, 90, 10982–10989 CrossRef CAS PubMed.
  23. X. Huang, Z. He, D. Guo, Y. Liu, J. Song, B. C. Yung, L. Lin, G. Yu, J. J. Zhu, Y. Xiong and X. Chen, Theranostics, 2018, 8, 3461–3473 CrossRef CAS PubMed.
  24. Y. K. Gao, N. Yang, T. T. You, C. M. Zhang and P. G. Yin, Sens. Actuators, B, 2018, 267, 129–135 CrossRef CAS.
  25. C. S. Fang, K. S. Kim, D. T. Ha, M. S. Kim and H. Yang, Anal. Chem., 2018, 90, 4776–4782 CrossRef CAS PubMed.
  26. G. Dutta and P. B. Lillehoj, Sci. Rep., 2018, 8, 17129 CrossRef PubMed.
  27. J. K. Ahn, H. Y. Kim, K. S. Park and H. G. Park, Anal. Chem., 2018, 90, 11340–11343 CrossRef CAS PubMed.
  28. J. C. Pyun, J. Jose and M. Park, Analyst, 2017, 142, 1720–1728 RSC.
  29. M. Kaisti, A. Kerko, E. Aarikka, P. Saviranta, Z. Boeva, T. Soukka and A. Lehmusvuori, Sci. Rep., 2017, 7, 15734 CrossRef PubMed.
  30. S. Park, J. E. Jeong, V. S. Le, J. Seo, B. Yu, D. Y. Kim, S. H. Kwon, S. Jon, H. Y. Woo and H. Yang, J. Am. Chem. Soc., 2018, 140, 2409–2412 CrossRef CAS PubMed.
  31. R. Huang, N. He and Z. Li, Biosens. Bioelectron., 2018, 109, 27–34 CrossRef CAS PubMed.
  32. Y. Wang, G. Zhao, Y. Zhang, B. Du and Q. Wei, Carbon, 2019, 146, 276–283 CrossRef CAS.
  33. Y. Zhang, A. Shoaib, J. Li, M. Ji, J. Liu, M. Xu, B. Tong, J. Zhang and Q. Wei, Biosens. Bioelectron., 2016, 79, 866–873 CrossRef CAS PubMed.
  34. M. S. Draz and H. Shafiee, Theranostics, 2018, 8, 1985–2017 CrossRef CAS PubMed.
  35. J. R. Choi, K. W. Yong, J. Y. Choi, A. Nilghaz, Y. Lin, J. Xu and X. Lu, Theranostics, 2018, 8, 1005–1026 CrossRef CAS PubMed.
  36. H. Zhou, J. Liu, J. J. Xu, S. S. Zhang and H. Y. Chen, Chem. Soc. Rev., 2018, 47, 1996–2019 RSC.
  37. J. Liu, Z. Geng, Z. Fan, J. Liu and H. Chen, Biosens. Bioelectron., 2019, 132, 17–37 CrossRef CAS PubMed.
  38. A. Salari and M. Thompson, Sens. Actuators, B, 2018, 255, 3601–3615 CrossRef CAS.
  39. Y. X. Chen, K. J. Huang and K. X. Niu, Biosens. Bioelectron., 2018, 99, 612–624 CrossRef CAS PubMed.
  40. J. Fan, D. Shang, B. Han, J. Song, H. Chen and J. M. Yang, Theranostics, 2018, 8, 5784–5800 CrossRef CAS PubMed.
  41. H. Xiang, Y. Wang, M. Wang, Y. Shao, Y. Jiao and Y. Zhu, Nanoscale, 2018, 10, 13572–13580 RSC.
  42. T. C. Liu, Y. C. Lee, C. Y. Ko, R. S. Liu, C. C. Ke, Y. C. Lo, P. S. Hong, C. Y. Chu, C. W. Chang, P. W. Wu, Y. Y. Chen and S. Y. Chen, Theranostics, 2018, 8, 4210–4225 CrossRef CAS PubMed.
  43. J. Xu, C. Yu, T. Feng, M. Liu, F. Li, Y. Wang and J. Xu, Nanoscale, 2018, 10, 19390–19398 RSC.
  44. E. A. Kwizera, R. O'Connor, V. Vinduska, M. Williams, E. R. Butch, S. E. Snyder, X. Chen and X. Huang, Theranostics, 2018, 8, 2722–2738 CrossRef CAS PubMed.
  45. F. Huang, L. Xue, H. Zhang, R. Guo, Y. Li, M. Liao, M. Wang and J. Lin, Theranostics, 2018, 8, 6263–6273 CrossRef CAS PubMed.
  46. X. Jin, J. Chen, X. Zeng, L. Xu, Y. Wu and F. Fu, Biosens. Bioelectron., 2019, 128, 45–51 CrossRef CAS PubMed.
  47. E. Stern, A. Vacic, N. K. Rajan, J. M. Criscione, J. Park, B. R. Ilic, D. J. Mooney, M. A. Reed and T. M. Fahmy, Nat. Nanotechnol., 2009, 5, 138 CrossRef PubMed.
  48. Y. Li, Y. Xiong, L. Fang, L. Jiang, H. Huang, J. Deng, W. Liang and J. Zheng, Theranostics, 2017, 7, 935–944 CrossRef CAS PubMed.
  49. W. Zhu, M. Saddam Khan, W. Cao, X. Sun, H. Ma, Y. Zhang and Q. Wei, Biosens. Bioelectron., 2018, 99, 346–352 CrossRef CAS PubMed.
  50. F. Gao, T. Fan, J. Wu, S. Liu, Y. Du, Y. Yao, F. Zhou, Y. Zhang, X. Liao and D. Geng, Biosens. Bioelectron., 2017, 96, 62–67 CrossRef CAS PubMed.
  51. C. C. Gu, P. P. Gai, X. J. Liu, J. Liu and F. Li, Sens. Actuators, B, 2018, 270, 1–8 CrossRef CAS.
  52. Y. H. Song, M. L. Xu, C. C. Gong, Y. Shen, L. Y. Wang, Y. Xie and L. Wang, Sens. Actuators, B, 2018, 257, 792–799 CrossRef CAS.
  53. D. Duan, H. Yang, Y. Ding, L. Li and G. Ma, Electrochim. Acta, 2019, 302, 137–144 CrossRef CAS.
  54. B. Chen and F. Wang, Nanoscale, 2018, 10, 19898–19905 RSC.
  55. L. Hua, Z. Hui, Y. Sun, X. Zhao, H. Xu, Y. Gong, R. Chen, C. Yu, J. Zhou, G. Sun and W. Huang, Nanoscale, 2018, 10, 21006–21012 RSC.
  56. R. Tian, Y. Li and J. Bai, Anal. Chim. Acta, 2019, 1058, 89–96 CrossRef CAS PubMed.
  57. I. R. Suhito, W. Lee, S. Baek, D. Lee, J. Min and T. H. Kim, Sens. Actuators, B, 2019, 288, 527–534 CrossRef CAS.
  58. S. Solovieva, M. Karnaukh, V. Panchuk, E. Andreev, L. Kartsova, E. Bessonova, A. Legin, P. Wang, H. Wan, I. Jahatspanian and D. Kirsanov, Sens. Actuators, B, 2019, 289, 42–47 CrossRef CAS.
  59. M. L. Moraes, L. R. Lima, J. C. Vicentini-Oliveira, A. V. G. de Souza, O. N. Oliveira, E. Deffune and S. J. L. Ribeiro, J. Nanosci. Nanotechnol., 2019, 19, 3772–3776 CrossRef PubMed.
  60. J. Y. Fu, X. S. An, Y. Yao, Y. M. Guo and X. Sun, Sens. Actuators, B, 2019, 287, 503–509 CrossRef CAS.
  61. X. Yu, L. Hu, F. Zhang, M. Wang, Z. Xia and W. Wei, Mikrochim. Acta, 2018, 185, 239 CrossRef PubMed.
  62. J. S. Park, S. W. Yi, H. J. Kim, H. J. Oh, J. S. Lee, M. Go, S. H. Shim and K. H. Park, Theranostics, 2018, 8, 5548–5561 CrossRef CAS PubMed.
  63. G. H. Pan, H. Wu, S. He, L. Zhang, Z. Hao, X. Zhang and J. Zhang, Nanoscale, 2018, 10, 23198 RSC.
  64. L. Wang, Y. Wen, L. Li, X. Yang, N. Jia, W. Li, J. Meng, M. Duan, X. Sun and G. Liu, Biosens. Bioelectron., 2018, 115, 91–96 CrossRef CAS PubMed.
  65. C. Y. Wang, Y. Qian, Y. Zhang, S. Y. Meng, S. Y. Wang, Y. H. Li and F. L. Gao, Sens. Actuators, B, 2017, 238, 434–440 CrossRef CAS.
  66. Y. Liu, E. Xiong, X. Li, J. Li, X. Zhang and J. Chen, Biosens. Bioelectron., 2017, 87, 970–975 CrossRef CAS PubMed.
  67. V. Mazanek, C. C. Mayorga-Martinez, D. Bousa, Z. Sofer and M. Pumera, Nanoscale, 2018, 10, 23149–23156 RSC.
  68. Y. Seok, H. A. Joung, J. Y. Byun, H. S. Jeon, S. J. Shin, S. Kim, Y. B. Shin, H. S. Han and M. G. Kim, Theranostics, 2017, 7, 2220–2230 CrossRef CAS PubMed.
  69. R. K. Daher, G. Stewart, M. Boissinot and M. G. Bergeron, Clin. Chem., 2016, 62, 947–958 CrossRef CAS PubMed.
  70. G. Dutta, S. Park, A. Singh, J. Seo, S. Kim and H. Yang, Anal. Chem., 2015, 87, 3574–3578 CrossRef CAS PubMed.
  71. P. Nandhakumar, B. Kim, N. S. Lee, Y. H. Yoon, K. Lee and H. Yang, Anal. Chem., 2018, 90, 807–813 CrossRef CAS PubMed.
  72. S. Park, G. Kim, J. Seo and H. Yang, Anal. Chem., 2016, 88, 11995–12000 CrossRef CAS PubMed.
  73. X. Huang, J. Song, B. C. Yung, X. Huang, Y. Xiong and X. Chen, Chem. Soc. Rev., 2018, 47, 2873–2920 RSC.
  74. T. Yang, R. Z. Yu, Y. H. Yan, H. Zeng, S. Z. Luo, N. Z. Liu, A. Morrin, X. L. Luo and W. H. Li, Sens. Actuators, B, 2018, 274, 501–516 CrossRef CAS.
  75. P. Wu, X. Hou, J. J. Xu and H. Y. Chen, Nanoscale, 2016, 8, 8427–8442 RSC.
  76. T. Yang, R. Yu, Y. Yan, H. Zeng, S. Luo, N. Liu, A. Morrin, X. Luo and W. Li, Sens. Actuators, B, 2018, 274, 501–516 CrossRef CAS.
  77. Z. Wang, R. Yu, H. Zeng, X. Wang, S. Luo, W. Li, X. Luo and T. Yang, Mikrochim. Acta, 2019, 186, 405 CrossRef PubMed.
  78. J. Zhang, L. L. Wang, M. F. Hou, Y. K. Xia, W. H. He, A. Yan, Y. P. Weng, L. P. Zeng and J. H. Chen, Biosens. Bioelectron., 2018, 102, 33–40 CrossRef CAS PubMed.
  79. H. Fang, J. Zhou and P. Jena, Nanoscale, 2018, 10, 22963–22969 RSC.
  80. Y. C. Wei, H. M. Ma, X. Ren, C. F. Ding, H. Wang, X. Sun, B. Du, Y. Zhang and Q. Wei, Sens. Actuators, B, 2018, 256, 504–511 CrossRef CAS.
  81. L. Y. Wang, M. L. Xu, Y. Xie, C. C. Qian, W. Ma, L. Wang and Y. H. Song, Sens. Actuators, B, 2019, 285, 264–270 CrossRef CAS.
  82. J. Fan, Q. Wei, E. J. Koay, Y. Liu, B. Ning, P. W. Bernard, N. Zhang, H. Han, M. H. Katz, Z. Zhao and Y. Hu, Theranostics, 2018, 8, 5986–5994 CrossRef CAS PubMed.
  83. L. Cui, M. Lu, Y. Li, B. Tang and C. Y. Zhang, Biosens. Bioelectron., 2018, 102, 87–93 CrossRef CAS PubMed.
  84. X. Li, B. T. Dou, R. Yuan and Y. Xiang, Sens. Actuators, B, 2019, 286, 191–197 CrossRef CAS.
  85. Y. H. Yuan, B. Z. Chi, S. H. Wen, R. P. Liang, Z. M. Li and J. D. Qiu, Biosens. Bioelectron., 2018, 102, 211–216 CrossRef CAS PubMed.
  86. X. H. Gao, R. J. Gui, H. J. Guo, Z. H. Wang and Q. Y. Liu, Sens. Actuators, B, 2019, 285, 201–208 CrossRef CAS.
  87. Y. H. Wang, G. Ning, H. Bi, Y. H. Wu, G. Q. Liu and Y. L. Zhao, Electrochim. Acta, 2018, 285, 120–127 CrossRef CAS.
  88. Y. Yu, C. Yu, T. Yin, S. Ou, X. Sun, X. Wen, L. Zhang, D. Tang and X. Yin, Biosens. Bioelectron., 2017, 87, 278–284 CrossRef CAS PubMed.
  89. D. Yao, W. Zhao, L. Zhang and Y. Tian, Analyst, 2017, 142, 4215–4220 RSC.
  90. L. Wang, C. C. Gong, Y. Shen, W. H. Ye, M. L. Xu and Y. H. Song, Sens. Actuators, B, 2017, 242, 625–631 CrossRef CAS.
  91. L. Li, Y. Zhang, L. N. Zhang, S. G. Ge, M. Yan and J. H. Yu, Sci. Bull., 2017, 62, 1114–1121 CrossRef CAS.
  92. X. H. Yu, Z. X. Wang, Y. F. Gao, F. Y. Kong, W. X. Lv, H. F. Ma and W. Wang, Int. J. Electrochem. Sci., 2018, 13, 2875–2886 CrossRef CAS.
  93. W. W. Zhao, X. D. Yu, J. J. Xu and H. Y. Chen, Nanoscale, 2016, 8, 17407–17414 RSC.
  94. X. H. Gao, R. J. Gui, K. Q. Xu, H. J. Guo, H. Jin and Z. H. Wang, New J. Chem., 2018, 42, 14796–14804 RSC.
  95. J. Liu, X. T. Jiang, R. Y. Zhang, Y. Zhang, L. M. Wu, W. Lu, J. Q. Li, Y. C. Li and H. Zhang, Adv. Funct. Mater., 2019, 29, 1807326 CrossRef.
  96. Z. Liu, H. Lin, M. Zhao, C. Dai, S. Zhang, W. Peng and Y. Chen, Theranostics, 2018, 8, 1648–1664 CrossRef CAS PubMed.
  97. X. Han, X. Jing, D. Yang, H. Lin, Z. Wang, H. Ran, P. Li and Y. Chen, Theranostics, 2018, 8, 4491–4508 CrossRef CAS PubMed.
  98. X. Yu, X. Cai, H. Cui, S. W. Lee, X. F. Yu and B. Liu, Nanoscale, 2017, 9, 17859–17864 RSC.
  99. S. Chen, Y. Xiang, M. K. Banks, C. Peng, W. Xu and R. Wu, Nanoscale, 2018, 10, 20043–20052 RSC.
  100. C. Zhu, M. Liu, X. Li, X. Zhang and J. Chen, Chem. Commun., 2018, 54, 10359–10362 RSC.
  101. Y. Lin, J. Wang, F. Luo, L. H. Guo, B. Qiu and Z. Y. Lin, Electrochim. Acta, 2018, 283, 798–805 CrossRef CAS.
  102. Y. Lin, J. Jia, R. Yang, D. Chen, J. Wang, F. Luo, L. Guo, B. Qiu and Z. Lin, Anal. Chem., 2019, 91, 3717–3724 CrossRef CAS PubMed.
  103. R. N. Ma, L. L. Wang, M. Zhang, L. P. Jia, W. Zhang, L. Shang, W. L. Jia and H. S. Wang, Sens. Actuators, B, 2018, 257, 678–684 CrossRef CAS.
  104. H. Li, N. Arroyo-Currás, D. Kang, F. Ricci and K. W. Plaxco, J. Am. Chem. Soc., 2016, 138, 15809–15812 CrossRef CAS PubMed.
  105. H. Li, J. Somerson, F. Xia and K. W. Plaxco, Anal. Chem., 2018, 90, 10641–10645 CrossRef CAS PubMed.
  106. J. Yu, H. Jin, R. Gui, Z. Wang and F. Ge, Talanta, 2017, 162, 435–439 CrossRef CAS PubMed.
  107. R. L. Ma, M. Jin and Y. Tian, J. Anal. Test., 2017, 1, 11 CrossRef.
  108. S. Chung, J. M. Moon, J. Choi, H. Hwang and Y. B. Shim, Biosens. Bioelectron., 2018, 117, 480–486 CrossRef CAS PubMed.
  109. D. Kim, H. J. Kwon, K. Shin, J. Kim, R. E. Yoo, S. H. Choi, M. Soh, T. Kang, S. I. Han and T. Hyeon, ACS Nano, 2017, 11, 8448–8455 CrossRef CAS PubMed.
  110. A. Barfidokht, S. Ciampi, E. Luais, N. Darwish and J. J. Gooding, Anal. Chem., 2013, 85, 1073–1080 CrossRef CAS PubMed.
  111. J. N. Chazalviel and P. Allongue, J. Am. Chem. Soc., 2011, 133, 762–764 CrossRef CAS PubMed.
  112. Y. F. Li, S. H. Ouyang, L. F. Tu, X. Wang, W. L. Yuan, G. E. Wang, Y. P. Wu, W. J. Duan, H. M. Yu, Z. Z. Fang, H. Kurihara, Y. Zhang and R. R. He, Theranostics, 2018, 8, 5713–5730 CrossRef CAS PubMed.
  113. X. Li, L. Jia, X. Chen, Y. Dong, X. Ren, Y. Dong, Y. Chen, L. Xie, M. Liu, C. Shiota, G. K. Gittes, L. Rui and Z. Chen, Theranostics, 2018, 8, 5960–5971 CrossRef CAS PubMed.
  114. W. Li, Y. Liu, Y. Wang, R. Li, P. Trang, W. Tang, Z. Yang, Y. Wang, X. Sun, X. Xing, S. Lu and F. Liu, Theranostics, 2018, 8, 5634–5644 CrossRef CAS PubMed.
  115. S. H. Bae, A. Jo, J. H. Park, C. W. Lim, Y. Choi, J. Oh, J. M. Park, T. Kong, R. Weissleder, H. Lee and J. Moon, Theranostics, 2019, 9, 1–10 CrossRef PubMed.
  116. J. Liu, Z. Geng, Z. Fan, J. Liu and H. Chen, Biosens. Bioelectron., 2019, 132, 17–37 CrossRef CAS PubMed.
  117. G. Gutierrez, H. Reines and M. E. Wulf-Gutierrez, Crit. Care, 2004, 8, 373 CrossRef PubMed.
  118. X. Wang, P. Gao, T. Yan, R. Li, R. Xu, Y. Zhang, B. Du and Q. Wei, Sens. Actuators, B, 2018, 258, 1–9 CrossRef CAS.
  119. Y. Lin, K. Liu, C. Wang, L. Li and Y. Liu, Anal. Chem., 2015, 87, 8047–8051 CrossRef CAS PubMed.
  120. H. Kaur, J. G. Bruno, A. Kumar and T. K. Sharma, Theranostics, 2018, 8, 4016–4032 CrossRef CAS PubMed.
  121. J. Noiphung, M. P. Nguyen, C. Punyadeera, Y. Wan, W. Laiwattanapaisal and C. S. Henry, Theranostics, 2018, 8, 3797–3807 CrossRef CAS PubMed.
  122. S. Liu, L. Fang, Y. Wang and L. Wang, Anal. Chem., 2017, 89, 3108–3115 CrossRef CAS PubMed.
  123. Y. Wu and R. Y. Lai, Anal. Chem., 2017, 89, 9984–9989 CrossRef CAS PubMed.
  124. K. Parate, C. Karunakaran and J. C. Claussen, Sens. Actuators, B, 2019, 287, 165–172 CrossRef CAS.
  125. H. Liang, Z. Zhou, R. Luo, M. Sang, B. Liu, M. Sun, W. Qu, F. Feng and W. Liu, Theranostics, 2018, 8, 5059–5071 CrossRef CAS PubMed.
  126. X. Liu, Z. Yan, Y. Sun, J. Ren and X. Qu, Chem. Commun., 2017, 53, 6215–6218 RSC.
  127. B. Meshram, A. Agrawal, S. Adil, S. Ranvir and K. Sande, Int. J. Curr. Microbiol. Appl. Sci., 2018, 7, 3305–3324 CrossRef.
  128. X. Zhuang, D. Chen, S. Wang, H. Liu and L. Chen, Sens. Actuators, B, 2017, 251, 185–191 CrossRef CAS.
  129. G. L. Xu, J. Z. Hou, Y. N. Zhao, J. Bao, M. Yang, H. B. Fa, Y. X. Yang, L. Li, D. Q. Huo and C. J. Hou, Sens. Actuators, B, 2019, 287, 428–436 CrossRef CAS.
  130. Z. Wang, B. Ma, C. Shen and L. Z. Cheong, Talanta, 2019, 197, 356–362 CrossRef CAS PubMed.
  131. F. Li, Z. Yu, X. Han, W. Shi, Y. Liu, H. Yan and G. Zhang, Sens. Actuators, B, 2018, 273, 480–487 CrossRef CAS.
  132. Y. Yan, P. Shi, W. Song and S. Bi, Theranostics, 2019, 9, 4047–4065 CrossRef PubMed.
  133. L. J. Zhao, R. J. Yu, W. Ma, H.-X. Han, H. Tian, R.-C. Qian and Y.-T. Long, Theranostics, 2017, 7, 876 CrossRef CAS PubMed.
  134. F. Canfarotta, J. Czulak, K. Betlem, A. Sachdeva, K. Eersels, B. van Grinsven, T. J. Cleij and M. Peeters, Nanoscale, 2018, 10, 2081–2089 RSC.
  135. Y. N. Zhang, Q. Niu, X. Gu, N. Yang and G. Zhao, Nanoscale, 2019, 11, 11992–12014 RSC.
  136. F. R. Caetano, E. A. Carneiro, D. Agustini, L. C. S. Figueiredo-Filho, C. E. Banks, M. F. Bergamini and L. H. Marcolino-Junior, Biosens. Bioelectron., 2018, 99, 382–388 CrossRef CAS PubMed.
  137. D. N. Pei, A. Y. Zhang, X. Q. Pan, Y. Si and H. Q. Yu, Anal. Chem., 2018, 90, 3165–3173 CrossRef CAS PubMed.
  138. Y. T. Kang, Y. J. Kim, J. Bu, Y. H. Cho, S. W. Han and B. I. Moon, Nanoscale, 2017, 9, 13495–13505 RSC.
  139. J. Ding, B. Li, L. Chen and W. Qin, Angew. Chem., Int. Ed., 2016, 55, 13033–13037 CrossRef CAS PubMed.
  140. D. Zang, L. Ge, M. Yan, X. Song and J. Yu, Chem. Commun., 2012, 48, 4683–4685 RSC.
  141. Z. Nie, C. A. Nijhuis, J. Gong, X. Chen, A. Kumachev, A. W. Martinez, M. Narovlyansky and G. M. Whitesides, Lab Chip, 2010, 10, 477–483 RSC.
  142. S. Wang, L. Wang, F. Li, B. Pingguan-Murphy, T. J. Lu and F. Xu, Biosens. Bioelectron., 2014, 54, 585–597 CrossRef PubMed.
  143. F. Arduini, S. Cinti, V. Caratelli, L. Amendola, G. Palleschi and D. Moscone, Biosens. Bioelectron., 2019, 126, 346–354 CrossRef CAS PubMed.
  144. J. Li, J. Xia, F. Zhang, Z. Wang and Q. Liu, Talanta, 2018, 181, 80–86 CrossRef CAS PubMed.

This journal is © The Royal Society of Chemistry 2019