Article pubs.acs.org/JPCC
Horizontally Aligned Carbon Nanotubes on a Quartz Substrate for Chemical and Biological Sensing Satoshi Okuda,† Shogo Okamoto, Yasuhide Ohno, Kenzo Maehashi,* Koichi Inoue, and Kazuhiko Matsumoto The Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka 567-0047, Japan ABSTRACT: We have developed electrolyte-gated sensors based on a fieldeffect transistor (FET) consisting of horizontally aligned single-walled carbon nanotubes (CNTs) synthesized on single-crystal quartz. Dense well-aligned CNTs serving as device channels enabled high current and large transconductance. Owing to these excellent device properties, the pH resolution was much better in the aligned-channel CNTFETs than in single-channel devices. For immunosensing, selective detection of human immunoglobulin E (IgE) by using aptamer-functionalized CNTFETs was demonstrated in the presence of nontarget proteins at much higher concentration. Moreover, measurements of sensor response versus IgE concentration produced data that fit well to the Langmuir adsorption isotherm. The developed sensors with aligned channels exhibited a drain current of 400-fold that of single-channel devices. Therefore, aligned-channel CNTFETs are useful for highly sensitive and practicable solution sensing.
1. INTRODUCTION Biosensors are becoming a vital part of modern life, from early diagnosis of life-threatening diseases to environmental monitoring to detection of allergens in food.1−4 Current biological sensing techniques commonly rely on an optical detection principle: fluorescence-based detection.5 Generally, that method is highly sensitive and specific, although a drawback is the necessity of laborious labeling to detect molecular recognition between a ligand and receptor. Recently, a number of studies have investigated electrochemical biodetection methods with the aim of developing label-free biosensors, which would be important in the move toward simplified testing, for example, with home-use devices. One approach to the electrical detection of biomolecules is to use nanoelectronic biosensors based on nanotubes and nanowires.6−8 In particular, carbon nanotube field-effect transistor (CNTFET)-based biosensors are a promising platform because CNTs have a number of unique and exotic properties, such as high carrier mobility, high aspect ratio, and excellent chemical stability.9−12 In previous studies, a single CNT or a few CNTs have been utilized as channels in CNTFETs.13−17 Such configurations pose several problems for the commercialization of biosensors. One is the small drain current (ID) of those CNTFETs. In an electrolyte, high bias cannot be applied between electrodes to prevent electrochemical reaction at their surfaces or to avoid leakage current. Consequently, the current that flows in a single CNT channel is limited to tens of nanoamperes at most, which is similar to the noise level in ID. A simple route to larger ID is to use a random network of CNTs as the channels of CNTFETs.18−24 This configuration offers several advantages: not only higher ID but also higher uniformity. The random © 2012 American Chemical Society
network configuration consists of both metallic and semiconducting CNTs. For the improvement of the signal-to-noise ratio (SNR), the larger transconductance (gm) is necessary because the larger transconductance leads to the large change in the drain current by adsorbing biomolecules with charges in solution. On the other hand, metallic CNT paths do not affect the transconductance. Thus, a number of semiconducting paths are a dominant factor of governing the gm, resulting in the improvement of sensitivity. However, Ishikawa et al. report that as the CNT density is increased the sensitivity of CNTFETs decreases.19 According to that report, within a 10 nm by 10 nm area around tube−tube junctions in a random network of CNTs, the electrical charge of biomolecules is strongly screened.19,25 This effect of a high-density CNT network impairs the sensing performance of CNTFETs. A high density of CNTs and a low density of tube−tube junctions are therefore needed for realizing highly sensitive CNTFET-based biosensors. Toward this end, controlling the growth direction of CNTs is necessary. Aligned arrays can avoid percolation transport, unusual scaling of device properties, and tube−tube junction resistance.26,27 However, there have been few studies that use aligned-CNT-based biosensors in the current literature.28 In recent years, controlled growth of CNTs has attracted much attention,29 and many techniques have been proposed to control the growth direction of CNTs by using, for example, an electric field,30 fast gas flow,31 growth on patterned SiO2 Received: February 16, 2012 Revised: August 13, 2012 Published: August 22, 2012 19490
dx.doi.org/10.1021/jp301542w | J. Phys. Chem. C 2012, 116, 19490−19495
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substrates with line and space grooves,32 and surface-guided growth on single-crystalline substrates such as quartz26,27,33−35 and sapphire.36−39 ST-cut quartz in particular is preferred for high-density packing of CNTs and large-scale production. On quartz substrates, CNTs are aligned along a specific crystal direction by angle-dependent van der Waals interactions between the CNTs and the quartz lattice. The density of the aligned CNTs synthesized on ST-cut quartz has reached as high as 60 CNTs/μm.35 In addition, CNTs grown on quartz have a narrow diameter distribution of 1.2 ± 0.3 nm with proper control of the growth conditions.35 In the present study, we utilized horizontally aligned CNTs grown on quartz substrates to develop high-performance CNTFET-based electrolyte-gated sensors. First, we investigated CNTFETs with multiple tubes serving as sensor transducers to detect changes in solution pH. Second, label-free real-time immunosensing was performed by using the proposed sensing platform. The biosensors exhibited specific detection of human immunoglobulin E (IgE), a class of antibodies that are associated with allergic disorders such as hay fever and atopic dermatitis.40,41 These results indicate that the CNTFETs with aligned CNTs have great promise for the development of chemical and biological sensors.
Figure 1. (a) Typical SEM image of horizontally aligned CNTs between catalyst lines synthesized on the quartz substrate. (b) Schematic illustration of the electrolyte-gated CNTFET sensor.
PBS), the antigen−antibody reaction usually occurs outside the electrical double layer, thus limiting the effect on the carriers in CNT channels. For this reason, the Debye length is an important factor in sensing performance.20 To overcome this constraint, an artificial oligonucleotide, IgE aptamers (∼2 nm; Famac Corp., Ltd.), were used as small receptors that fall within the Debye length. Compared with antibodies, the aptamers have advantages in highly sensitive biosensing: not only smaller size but also lower cost and higher stability.45 In this report, IgE aptamer DNA oligonucleotides (D17.4ext) with 5′-amino modification were custom-synthesized, and the base sequence was 5′-NH2 GCG CGG GGC ACG TTT ATC CGT CCC TCC TAG TGG CGT GCC CCG CGC-3′. Figure 2 shows a
2. EXPERIMENTAL METHODS Fabrication of CNTFET with Horizontally Aligned CNTs. CNTFETs were fabricated by position-controlled growth42 on quartz substrates. First, ST-cut single-crystal quartz (Kyocera Kinseki Corp.) was annealed at 900 °C for 8 h prior to the CNT growth. Lines of thin-film iron catalyst (0.5 nm) were patterned by photolithography and electron-beam evaporation onto the substrate. The lines were oriented perpendicular to the X axis of the quartz crystal, which corresponded to the direction of the aligned CNTs. After a liftoff process, samples were annealed at 550 °C for 1 h to form isolated iron oxide nanoparticles. CNTs were grown by thermal chemical vapor deposition (CVD) using ethanol at 900 °C for 20 min. Raman spectroscopy confirmed that single-walled CNTs were synthesized.43 The scanning electron microscopy (SEM; S-4800, Hitachi) image presented in Figure 1a shows typical horizontally well-aligned single-walled CNTs between lines of iron catalyst. After assembly of the CNT array, source and drain electrodes (Au 30 nm/Ti 2 nm) were formed in the regions between catalyst lines on the CNT/quartz substrate by a photolithography and liftoff process. The channel length and width were approximately 3 and 100 μm, respectively. Electrical Measurements in Electrolyte and Preparation of Immunosensor. Figure 1b shows a schematic diagram of the sensor architecture that used the CNTFET with aligned CNTs as a transducer. To utilize the CNTFET in solution, a silicone rubber barrier was attached to the device. Electrical measurements were carried out by using a semiconductor device analyzer (B1500A, Agilent). A gate voltage was applied via a Ag/AgCl reference electrode (Cypress Systems), which is commonly used in electrochemical sensing to minimize the perturbation from adding target species into solution.14 To enable the selective recognition and binding of IgE (Yamasa Corp.), the surface of CNTs should be functionalized with specific receptors.15,44 A general approach to detect specific biomolecules is to use antigen−antibody reactions. However, antibody-functionalized CNTFET sensors are not suitable for highly sensitive biosensing. Since a typical antibody size (∼15 nm) exceeds the Debye length in solution (3 nm in 10 mM
Figure 2. Procedure to immobilize IgE aptamers on CNTs. (a) Incubation of a CNT with methanol solution of 1-pyrenebutanoic acid succinimidyl ester, (b) immobilization of 1-pyrenebutanoic acid succinimidyl ester on the CNT, and (c) functionalization of aptamers with the CNT.
procedure to immobilize IgE aptamers on the CNTs. Before functionalization with aptamers, the device was incubated with 5 mM methanol solution of 1-pyrenebutanoic acid succinimidyl ester (Life Technologies Corp.), as shown in Figure 2a, which was used as a bifunctional linker.46 The pyrenyl group of the linker strongly interacts with the six-membered rings of CNTs via π-stacking, as shown in Figure 2b. The succinimidyl ester of the linker, which is anchored onto CNTs, reacts with the amino 19491
dx.doi.org/10.1021/jp301542w | J. Phys. Chem. C 2012, 116, 19490−19495
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agents. In electrolyte, CNTFETs with a single channel often exhibit large current fluctuation under a low bias. In a previous study, alternating current measurements were carried out to decrease unintentional noise around the CNTFETs.16 Other researchers have made effective use of noisy environments to enhance the sensitivity of CNTFETs through stochastic resonance.13 Here, to investigate the fluctuation of ID, the time course of ID for aligned-channel CNTFETs was measured. For comparison, the ID fluctuation of a single-channel CNTFET was also measured. The real-time response recorded by tracking ID in phosphate buffer is shown in Figure 3b. The blue and red lines correspond to ID of the aligned- and singlechannel CNTFETs, respectively, ID was monitored for 10 min and was normalized by the average of ID. Compared with the single-channel CNTFET, the aligned-channel CNTFET exhibited much lower ID fluctuation. The residual standard deviation (RSD) was calculated as 2.4% and 0.05% for the single- and aligned-channel CNTFETs, respectively, owing to the significant increase in ID. Therefore, the multichannel configuration will contribute to improved sensing performance. Subsequently, we investigated the reproducibility of the electrical characteristics of aligned-channel CNTFETs. Histograms showing the distribution of ID at VEG = −0.2 V and VD = 0.2 V are plotted for the aligned- and single-channel CNTFETs (52 devices each) in Figure 4. In the case of the aligned-channel CNTFETs, ID had a normal distribution, and the average of ID
groups of the aptamers to form amide bonds for aptamer functionalization, as shown in Figure 2c. Linkers that failed to bind to the aptamers were inactivated by reaction with ethanolamine (10 mM) for 1 h. To demonstrate the selective detection of IgE, bovine serum albumin (BSA; Sigma-Aldrich Inc.) and avidin (MP Biomedicals, LLC) were used as the nontarget proteins.
3. RESULTS AND DISCUSSION 3.1. Characterization of CNTFET on the Quartz Substrate in Electrolyte. Figure 3a shows the representative
Figure 3. (a) ID versus VG at VD = 0.2 V in back-gate operation in air (blue) and electrolyte-gate operation in phosphate buffer (red). Inset shows an expanded plot from VG = −40 to 40 V. (b) Normalized conductance versus time in phosphate buffer for the single-channel (red) and aligned-channel (blue) CNTFETs.
transfer characteristics of back-gate operation in air and electrolyte-gate operation in 10 mM phosphate buffer (pH 7.4) at a drain voltage (VD) of 0.2 V. In air, ID was barely affected by changes in back-gate voltage (−0.2 to 0.2 V) because quartz was used as the back gate and the presence of metallic CNTs. In phosphate buffer, on the other hand, ID was substantially modulated by the electrolyte-gate voltage (VEG) from the reference electrode. The CNTFET exhibited p-type semiconductive behavior in phosphate buffer. The inset shows the plot from VG = −40 to 40 V. The gm of the CNTFET was estimated to be 0.02 and 144 μS in air and phosphate buffer, respectively. In phosphate buffer, gm was markedly higher since the electrical double layer on the CNT surface acts as an extremely thin top-gate insulator. The higher gm of the CNTFET in phosphate buffer is a nontrivial factor in solution sensing because large gm leads to a large ID change due to binding of the target analyte. The fluctuation of ID also affects the detection sensitivity because the fluctuation easily obscures true signals from target
Figure 4. Distribution of ID in phosphate buffer at VD = 0.2 V and VEG = −0.2 V for (a) aligned-channel CNTFET on the quartz substrate and (b) single-channel CNTFET on the Si/SiO2 substrate. For each histogram, 52 devices were measured. 19492
dx.doi.org/10.1021/jp301542w | J. Phys. Chem. C 2012, 116, 19490−19495
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and RSD was estimated as 311 μA and 12.8%, respectively (Figure 4a). Furthermore, all aligned-channel devices operated as FETs. In contrast, ID of single-channel devices had a random distribution because their electrical characteristics were strongly affected by the characteristics of individual CNTs (Figure 4b). The single-channel devices exhibit 226% of RSD, which was about 20-fold wider ID distribution compared with that of aligned-channel devices. Therefore, CNTFETs constructed of multiple channels have good device-to-device uniformity, which is an essential trait for commercialization of CNTFET-based electrolyte-gated sensors. 3.2. Sensing Performance of an Aligned-Channel CNTFET-Based Sensor. The pH dependence of the electrical characteristics of aligned-channel CNTFETs was evaluated. Figure 5a shows the time course of ID monitored in a series of
devices. Although the on/off ratio of ID in our device was small (
