KR101786366B1 - Method for real-time detection of silver nanowire using quartz crystal microbalance and thio-silver bond - Google Patents

Abstract

The present invention relates to a method of detecting nanowires in a sample using a quartz vibration scale and thiol-silver bonding, and more particularly,
a) combining the silver nanowire and the linker DNA by mixing the linker DNA in a sample containing the nanowire;
b) binding the probe DNA onto the quartz electrode;
c) performing hybridization of the linker DNA and the probe DNA by flowing the sample into a flow cell including the crystal electrode; And
d) analyzing that the resonance frequency of the quartz electrode is shifted as the weight of the quartz electrode is increased by the hybridization of step c)
A method for detecting nanowires in real time.
According to the present invention, it is possible to detect the nanowires in an unlabeled, high-sensitivity, direct, and real-time manner in a sample, and detect silver nanowires with a very precise detection limit for various samples including deionized water and drinking water , And it is also possible to detect only the silver nanowires among other one-dimensional nanomaterials with very high selectivity.

Description

TECHNICAL FIELD The present invention relates to a method for detecting nanowires in a sample using quartz crystal balance and thiol-silver bond,

The present invention relates to a method for detecting nanowires in a sample using quartz vibration scales and thiol-silver bonds.

Recently, research on silver nanomaterials (AgNMs) has been actively carried out, which shows that silver nanomaterials have unique physical, chemical, electrical and optical properties, which allows a variety of products, Products. Thus, the annual production of silver nanomaterials is steadily increasing, and is now reported to be about 10,000 tonnes per year.

Among the various silver nanomaterials, silver nanowires (AgNWs) have received much attention because AgNWs have high aspect ratios to diameters of less than 100 nm and as much as 50 μm. In addition, AgNWs exhibit higher electrical and thermal conductivity than other types of nanowires and exhibit strong surface plasmon resonance. Thus, AgNWs are widely used in a variety of applications, such as touch screens, solar cells, electrochemical sensors,

Although AgNWs are rapidly applied to a variety of fields, a number of documents have been reported on the effect of AgNWs on living organisms. AgNWs released in the environment during the production, use and disposal of products containing AgNWs are absorbed by humans either directly or through drinking water or indirectly through the food chain. Since AgNWs are small in size and high in aspect ratio, AgNWs are highly reactive and considered to have a high ability to penetrate into living tissues. In this case, they exhibit cytotoxic effects similar to those caused by carbon nanotubes It is known to be able to.

Recently, AgNWs have been reported for toxicity, and Scanlan et al. Have reported on the cytotoxicity of four classes of AgNWs on daphnia. Specifically, when daphnia is exposed to a growth medium containing AgNWs, AgNWs and Ag-rich precipitates are observed within the hemolymph, demonstrating toxicity of AgNWs to daphnia. Kim et al. Have also studied the toxicity of AgNWs to human erythrocytes. When human erythroid cells are exposed to AgNWs-containing solutions, agglutinates of AgNWs are observed in the form of spicules on the cell membrane , Which also implies a toxic effect of AgNWs on humans. In addition, there are references to various toxic effects of AgNWs on various organisms.

On the other hand, a quartz crystal microbalance is a measuring device using quartz crystal microbalance. The quartz crystal microbalance changes the resonant frequency according to a mass change of a substance bonded to a surface of a quartz crystal, Device. Conventionally, there has been disclosed a technique using a quartz oscillating microbalance including a biomolecule having a predetermined functional group covalently bonded to silver for detecting an antigen (Patent Document 1), and further, by using DNA for probe, (Patent Document 2) discloses a quartz oscillator microbalance capable of detecting a quartz crystal oscillator (Patent Document 2).

However, until now, no technology has been reported utilizing a specially designed quartz vibration scale for the detection of nanowires.

Patent Document 1: Japanese Patent Publication No. 4234596 Patent Document 2: Korean Patent Laid-Open No. 10-2009-0039456

Accordingly, it is an object of the present invention to solve the above-mentioned problems of the prior art, and it is an object of the present invention to provide a silver nanowire which is very toxic to the human body and environment and can be detected by a simple and economical method at a high sensitivity and in real time, A method for real-time detection of silver nanowires capable of detecting only nanowires with very high selectivity.

Accordingly, in order to solve the above problems,

a) combining the silver nanowire and the linker DNA by mixing the linker DNA in a sample containing the nanowire;

b) binding the probe DNA onto the quartz electrode;

c) performing hybridization of the linker DNA and the probe DNA by flowing the sample into a flow cell including the crystal electrode; And

d) analyzing that the resonance frequency of the quartz electrode is shifted as the weight of the quartz electrode is increased by the hybridization of step c)

The present invention provides a method for real-time detection of nanowires.

According to an embodiment of the present invention, the step a) may be performed by mixing the linker DNA with a reducing agent to activate the thiol group of the linker DNA, and then binding the activated thiol group to the silver nanowire.

According to another embodiment of the present invention, the reducing agent may be selected from the group consisting of tris (2-carboxyethyl) phosphine, dithiothreitol, dithiolbutylamine, beta-mercaptoethanol and mixtures thereof.

According to another embodiment of the present invention, the resonant frequency shift of step d) may be calculated by the following equation:

&Quot; (1) "

Where f ₀ is the natural frequency of the non-hybridized quartz electrode, A is the active area of the quartz electrode, and ρ _q and μ _q are the density and shear modulus of the quartz electrode, respectively.

According to another embodiment of the present invention, the coupling of the probe DNA to the quartz electrode in step b) may be performed by observing the surface of the quartz crystal electrode by an atomic force microscope, Can be confirmed by calculating the surface roughness of the quartz crystal electrode surface.

According to another embodiment of the present invention, the binding of the silver nanowire and the linker DNA in step a) can be confirmed by analyzing the UV-Visible absorption spectrum of the sample.

According to another embodiment of the present invention, the method of detecting nanowires in the sample may have a detection limit of 1 ng / ml.

According to the present invention, it is possible to detect the nanowires in an unlabeled, high-sensitivity, direct, and real-time manner in a sample, and detect silver nanowires with a very precise detection limit for various samples including deionized water and drinking water , And it is also possible to detect only the silver nanowires among other one-dimensional nanomaterials with very high selectivity.

1 is a schematic overview of a method for detecting silver nanowires according to the present invention.
2A and 2B are AFM images of the pure electrode 2a and the DNA-immobilized electrode 2b, and FIG. 2C is a graph showing a comparison of surface roughness of the pure electrode and the DNA-immobilized electrode.
3A and 3B are atomic force microscope images of gold-attached silver nanowires 3a and pure silver nanowires 3b.
4 is a graph showing the UV-visible absorption spectrum of silver nanowires (red lines) and DNA-silver nanowires (black lines).
5 is a diagram showing an atomic force microscope image 5a, an SEM image 5b, and an EDX spectrum 5c of a quartz crystal electrode after silver nanowire detection is completed.
Figures 6a and 6b illustrate real time resonance frequency shifts measured using the silver nanowire detection method according to the present invention for various silver nanowire concentration conditions and control (black line) in deionized water, And a graph (6b) showing the average frequency shift at the time of the elapse of time (the control group is a gray bar).
7 is a graph comparing resonance frequency shift values for silver nanowires with resonance frequency shifts for CNTs and SiO ₂ nanowires.
Figures 8a and 8b show the real time resonance frequency shifts measured using the silver nanowire detection method according to the present invention for various silver nanowire concentration conditions and the control group (black line) (6b) showing the average frequency shift at the elapsed time point (the control group is a gray bar).

Hereinafter, the present invention will be described in more detail.

The present invention seeks to provide an unlabeled, highly selective, direct and real-time detection method for silver nanowires using Quartz Crystal Microbalance and DNA.

FIG. 1 is a schematic diagram of a method for detecting silver nanowires according to the present invention. Referring to FIG. 1,

a) combining the silver nanowire and the linker DNA by mixing the linker DNA in a sample containing the nanowire;

b) binding the probe DNA onto the quartz electrode;

c) performing hybridization of the linker DNA and the probe DNA by flowing the sample into a flow cell including the crystal electrode; And

d) analyzing that the resonance frequency of the quartz electrode is shifted as the weight of the quartz electrode is increased by the hybridization of step c)

.

The binding of the silver nanowire and the linker DNA in the sample during step a) may be performed by mixing the linker DNA with a reducing agent to activate the thiol group of the linker DNA and then binding the activated thiol group to the silver nanowire The activated thiol group is known to bind well to metal surfaces such as gold and silver.

The reducing agent may be selected from the group consisting of, but not limited to, tris (2-carboxyethyl) phosphine, dithiothreitol, dithiolbutylamine, beta-mercaptoethanol and mixtures thereof .

In step (b), the probe DNA is hybridized with the linker DNA on the quartz crystal electrode separately from the step (a). The step (b) , Followed by washing and drying.

As a next step, a step of performing hybridization of the linker DNA bound to the silver nanowire and the probe DNA bound to the crystal electrode is performed (step (c)), and the linker DNA and the probe DNA They have a complementary sequence to form double stranded DNA by hybridization.

In the method according to the present invention, the nanowire is detected in the sample using the phenomenon that the resonance frequency of the electrode is shifted due to the increase of the weight of the quartz electrode due to the DNA hybridization. The frequency shift can be calculated by the following equation:

&Quot; (1) "

Where f ₀ is the natural frequency of the non-hybridized quartz electrode, A is the active area of the quartz electrode, and ρ _q and μ _q are the density and shear modulus of the quartz electrode, respectively.

Meanwhile, in steps a) and b), it may be additionally performed to check whether the linker DNA and the probe DNA are well bound to the silver nanowire and the quartz electrode, respectively. For example, The binding of the probe DNA to the quartz electrode can be confirmed by observing the surface of the quartz crystal electrode by an atomic force microscope or by calculating the surface roughness of the quartz crystal electrode surface appearing in the image obtained by the atomic force microscope The binding between the silver nanowire and the linker DNA of step a) can be confirmed by analyzing the UV-Visible absorption spectrum of the sample.

As can be seen from the data of the following examples, the present invention can detect only nanowires in a sample with excellent selectivity even in the presence of various other interfering ions in the sample, and its detection limit (detection of sensitivity) It is also very good at 1 ng / ml.

EXAMPLES Hereinafter, the present invention will be described in more detail with reference to Examples. However, the following Examples are intended to assist the understanding of the present invention and are not intended to limit the scope of the present invention.

Materials and materials

Tris-EDTA buffer and tris (2-carboxyethyl) phosphine hydrochloride (TCEP) were purchased from Sigma Aldrich. The following DNAs were purchased from Integrated DNA Technology (CA, USA): 5'- / 5ThioMC6-D / TTG AAG ATG CTG GTA GAA GA-3 ' ATC TTC AA-3 '(Linker DNA). Chrome / gold crystals were purchased from Stanford Research Systems (CA, USA) and commercial TESP cantilever tips were used for all imaging experiments (Bruker, Santa Barbara, CA, USA).

Device

SRS QCM 200 was purchased from Stanford Research Systems. An Innova atomic force microscope equipped with a Nanodrive controller from Bruker was used to analyze the surface of the quartz electrode. To confirm the detection of AgNWs, S-4800 field emission scanning electron microscope (Hitachi, Japan) was used. Solution flow was generated using a Ne-1000 syringe pump (New Era Pump Systems, Inc.). A DR-4000U (Hach company, USA) spectrophotometer was used to confirm the attachment of DNA on AgNWs.

AgNWs  Produce

AgNWs were prepared by solvent thermo - synthesis. First, the flask was cleanly washed with aqua regia (HCl / HNO3 ³ : 1 v / v), rinsed with ethyl alcohol, and then dried under a nitrogen flow. It was then added to a polyvinyl pyrrolidone (PVP) of 0.5 g KNO ₃ and 2 g of in 100 ml of ethylene glycol (EG). The mixture was stirred at 433 K at 260 rpm for 1 hour, 4 ml of 3 mM HCl solution was added to the mixed solution and 20 ml of 94 mM AgNO ³ was added. Finally, AgNWs were prepared after heating the mixture at 433 K for 1 hour.

Surface modification of the quartz electrode

The quartz electrode was washed with triple distilled water, ethyl alcohol, and acetone, and then dried in a desiccator. The electrode was then immersed in a solution containing 10 [mu] M probe DNA and 30 mM Tris-EDTA buffer solution. Thereafter, the quartz electrode was rinsed with deionized water and dried once again in decahydrate.

Linker DNA and Ag Of NWs  join

1.5 μl of a solution containing 10 mM TECP (Tris (2-carboxyethyl) phosphine) was mixed with 1 ml of a solution containing 10 μM linker DNA and left for 1 hour to activate the thiol group. The solution was then added to a 4 ml solution containing AgNWs to conjugate DNA and AgNWs. For a smooth coupling, the solution was sonicated for 2 hours using a tip sonicator.

Characterization

Characterization of the QCM quartz electrode was performed using a tapping mode atomic force microscope (AFM) to analyze the surface. All surface images were acquired for a 3 ㎛ x 3 ㎛ area with a scan speed of 0.65 Hz. The experiments were carried out at room temperature. Heights and surface images were analyzed using Nanoscope software V1.20 (Bruker, Santa Barbara, CA, USA). In addition, scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX) techniques were used to characterize the quartz electrode surface.

QCM  analysis

AgNWs detection was performed using a chromium / gold quartz crystal electrode with a fundamental frequency of 5 MHz. The sampling period was 10 s, and the detection resolution was 0.01 Hz. Linker DNA was conjugated to various concentrations of AgNWs (10 μg / ml, 1 μg / ml, 100 ng / ml, 10 ng / ml, 1 ng / ml, 100 μg / ml) to calculate detection sensitivity. The solution was supplied by a syringe pump through a QCM flow cell at a flow rate of 7 ml / h. The resonance frequency shift is related to the mass change, which is termed Sauerbrey's equation:

Where f ₀ is the natural frequency of the unsupported electrode, A is the active area of the electrode, and ρ _q and μ _q are the density and shear modulus of the electrode, respectively.

Drinking water AgNWs  detection

To assess the efficiency of AgNWs detection in real-life applications, detection of AgNWs in ready-made drinking water (Dongwon Minnemain, Korea) was performed. Prior to sampling, the drinking water was filtered using a syringe filter (0.2 μm pore size, Minisart, Sigma-Aldrich). Samples were prepared similar to deionized water samples, except that ready-made drinking water was used instead of deionized water. All drinking water samples were analyzed by the same method used for deionized water samples.

AgNWs  Selection detection

Selective detection of AgNWs was investigated using single-walled carbon nanotubes (SWNTs) and silicon oxide nanowires (SiNWs), which are widely used as one-dimensional materials. The concentrations of the two substances were fixed at 10 μg / ml, which is the highest concentration in the AgNWs detection method. All samples were prepared similarly to the AgNWs samples and the resonance frequency shift obtained from the selectivity experiments was compared to the values in the AgNWs experiments.

Results and review

Ag NWs  Detection mechanism

In the present invention QCM and DNAs are used to provide unlabeled, highly selective, direct and real-time detection methods for AgNWs. FIG. 1 shows a schematic outline of a method of detecting silver nanowires according to the present invention.

Referring to FIG. 1, one DNA (linker DNA) was introduced to modify AgNWs to a specific DNA sequence, and other DNA (probe DNA) was functionalized on a quartz electrode. When the solution of the linker DNA-modified AgNWs (DNA-AgNWs) flows into the flow cell containing the quartz electrode, the DNA-AgNWs bind to the quartz electrode through DNA hybridization, This is because it is designed so as to be hybridized. Thus, the total weight of the quartz crystal electrode increases, and the resonant frequency of the electrode shifts according to the Sauerbrey equation. AgNWs detection is performed in real time by analyzing this resonance frequency shift.

Probe  DNA fixation confirmation

Prior to the detection of AgNWs, in the present invention, probe DNA was immobilized on a chromium / gold correction electrode using a thiol-gold bond. AFM images were obtained for pure electrodes and DNA-immobilized electrodes (FIGS. 2A and 2B) to confirm whether DNA immobilization was successfully performed on the electrodes.

As shown in Fig. 2A, a flat, spot-free surface was observed for the pure electrode. However, as shown in Figure 2b, a rough surface was observed for the DNA-immobilized electrode. Surface roughness was also calculated from the AFM images and the roughness was calculated from the arithmetic mean of Ra calculated using the following equation:

Illumination data includes n- order points along the trace and y _i is the vertical distance from the mean to the i-th data point. Ra values for the pure electrode and the DNA-immobilized electrode were 0.64 +/- 0.05 and 1.14 +/- 0.15 nm, respectively (see Figure 1c). Increased surface roughness is another evidence that the probe DNA was immobilized on the electrode.

DNA- AgNWs  Confirm

To confirm the junction between the linker DNA and AgNWs, probe DNA (DNA-AuNPs) bound to gold nanoparticles was introduced. Because thiol groups easily bind to gold particles, thiolated probe DNAs attach to gold nanoparticles. As a control, DNA-AuNPs were introduced into pure AgNWs and DNA-AuNPs were not observed on AgNWs (see FIG. 3a). However, when DNA-AuNPs were introduced into DNA-AgNWs, DNA-AuNPs were observed on DNA-AgNWs, which is due to DNA hybridization between probe DNA and linker DNA (see FIG. In the present invention, a UV-Visible absorption spectrum was obtained to confirm the DNA-AgNWs junction. The spectra were obtained for pure AgNWs and DNA-AgNWs in deionized water. For the pure AgNWs absorption spectrum, two distinct absorption peaks were observed at about 350 and 390 nm (see FIG. 4). The peak at 350 nm corresponds to the surface plasmon resonance (SPR) of AgNWs, which is similar to bulk silver. In addition, the peak at 390 nm is due to the transversal SPR mode of AgNWs. However, the peak of DNA-AgNWs shifted slightly from 390 nm to 398 nm of pure AgNWs. This absorption shift represents evidence that the linker DNAs are conjugated onto AgNWs. These results confirm that the linker DNAs are conjugated with AgNWs.

AgNWs  detection

In order to confirm the sensitivity of the detection method according to the present invention, the frequency shift of the quartz electrode was measured under various conditions of AgNWs. A control experiment was performed to confirm the interaction of DNA-AgNWs with the pure electrode. As a result, no frequency shift was observed when 1 ㎍ / ml AgNWs solution was exposed to the pure electrode. Furthermore, no AgNWs were observed in AFM images for the pure electrode after flowing the DNA-AgNWs solution (Figure 5). Thus, it can be seen that AgNWs do not interact with the pure electrode.

For AgNWs detection, QCM was first stabilized with deionized water. The flow was then converted to AgNWs solution. The frequency shift was not observed for the initial 20 minutes because the AgNWs solution took time to reach the electrode. However, a remarkable frequency shift was observed after 20 minutes (see Fig. 6A). When the AgNWs solution reaches the electrode in the flow cell, DNA-AgNWs hybridize with the probe DNA of the electrode. As a result, the total weight of the electrode increases, and a frequency shift occurs. The frequency shift continuously increases with time, and a higher frequency shift value is observed at higher AgNWs concentrations. Figure 6b summarizes the frequency shift response with AgNWs concentration after 60 minutes of detection. Specifically, when the concentrations of AgNWs were 10 ⁴ , 10 ³ , 10 ² , 10 ¹ , 10 ⁰ , 10 ^-1 , and 0 (control group), each frequency shift was 10.12 ± 0.86, 5.51 ± 0.25, 3.79 ± 0.76, 2.39 ± 0.38, 1.46 ± 0.26, 0.52 ± 0.20, and 0.31 ± 0.38 Hz, respectively. As the AgNWs concentration decreases, the frequency shift also decreases. The lowest concentration of 10 ^-1 ng / ml showed a similar response to the control (0 ng / ml), but the frequency shift measured at 10 ⁰ ng / ml was about 4.7 times It was a large value, which means that the limit of detection (LOD) of the method according to the invention 10 ⁰ ng / ml.

AgNWs  Detection confirmation

In order to confirm the detection of AgNWs, the surface of the electrode was analyzed using AFM, SEM, and EDX. As shown in Figure 5, AgNWs structures were observed on the electrode after detection. As a result of the point analysis using EDX, three kinds of elements (Au, Si, and Ag) were confirmed on the electrode (Fig. 5). As shown in Table 1 below, the weight percent and atomic percentages of silver were 43.74 and 56.46, respectively.

Significant silver peaks indicate that the observed structures correspond to AgNWs. These results confirm the detection of AgNWs. Au (55.38 wt%, 39.15 at%) and Si (0.89 wt%, 4.40 at%) were also observed on the electrode because the original composition of the electrode contains gold and silica.

Detection selectivity

Further, the detection selectivity of the method according to the present invention was evaluated because selectivity is an important criterion for determining the reliability of the detection method. To test the detection selectivity, other one-dimensional nanomaterials including single-walled carbon nanotubes (SWNTs) and silicon oxide nanowires (SiNWs) were used. SiNWs have been developed for biotechnological applications and are known as one-dimensional nanomaterials that exhibit cytotoxicity. SWNTs are also widely used nanomaterials. To compare resonance frequency shifts for SWNTs, SiNWs, and AgNWs, the relative frequency shift was defined as:

% N = 100 x (? _{F m} /? _{F AgNWs} )

In the above equation ,? F _m And? F _{Ag NWs} represent the frequency shift of 1-dimensional nanomaterials at 10 μg / ml concentration and the average frequency shift of AgNWs at 10 μg / ml concentration, respectively. As shown in Fig. 7, the relative frequency shifts of SWNTs and SiNWs were 11.03 ± 0.56% and 11.69 ± 2.60%, respectively. Although linker DNA can surround SWNTs by a pi-pi stack, binding between probe DNA and linker DNA can alter the structure of the linker DNA and interfere with the interaction between SWNTs and linker DNA (A Graphene Platform for Sensing Biomolecules). This interaction releases SWNTs from the linker DNA, and thus the SWNTs can not be adsorbed on the QCM electrode. In the case of SiNWs, thiol groups do not bond to SiO ₂ . Thus, the selectivity of SWNTs and SiNWs is negligible compared to AgNWs, which is indicative of the selectivity for the AgNWs of the present invention.

Applicability to drinking water

Detection of AgNWs in ready-made drinking water (Dongwon Minnemain, Korea) was performed to assess whether the method according to the present invention could be applied in real life. The potable water used in this experiment is a large amount of ^{Mg 2 + (0.54 mM),} Ca 2 + (5.21 mM), Na + (1.74 mM), F - and contained the (0.05 mM), and K ⁺ (0.13 mM) ion . The sample preparation was performed in the same manner as the deionized water sample, except that ready-made drinking water was used instead of deionized water. Figure 8a shows the actual frequency shift under various concentrations of AgNWs. The measured frequency shift values for the samples with concentrations of 10 ⁴ , 10 ³ , 10 ² , 10 ¹ , 10 ⁰ , 10 ^-1 , and 0 (control group) ng / ml were 10.55 ± 0.83, 5.81 ± 0.51, 3.85 ± 0.85, 2.73 ± 0.72, 1.13 ± 0.59, 0.51 ± 0.11, and 0.41 ± 0.10 Hz, respectively (FIG. A frequency shift was observed after conversion to AgNWs solution. Due to the high selectivity of the detection mechanism of the process according to the invention, the results obtained for drinking and deionized water were not significantly different. Despite the presence of interfering ions in the drinking water, the measured detection limit was 100 ng / ml. Sensitivity as well as frequency shifts in drinking water were also similar to those in deionized water. Although metal ions have been shown to interfere with the hybridization between linker DNA and probe DNA, DNA hybridization has also been strongly stabilized by metal ions. Thus, the frequency shift of the drinking water was similar to the frequency shift of the deionized water.

Taken together, the present invention first provides unlabeled, highly sensitive, direct and real-time detection methods for AgNWs. In the present invention, the detection is performed based on the resonance frequency shift of the QCM according to the two DNAs and the reaction. When the probe-DNA-immobilized electrode is exposed to a solution of DNA-AgNWs, the DNA-AgNWs hybridize with the probe DNAs on the electrode to enable the detection of AgNWs. By using the method according to the present invention, AgNWs could be detected with a detection limit of 1 ng / ml for both deionized and potable water. Furthermore, among the other one-dimensional nanomaterials, AgNWs could be selectively detected, demonstrating the excellent selectivity of the present invention. Thus, the method according to the present invention can be viewed as providing a means of real-time monitoring for detecting and analyzing the toxicity of AgNWs present in a real environment.

Claims (7)

a) combining the silver nanowire and the linker DNA by mixing the linker DNA in a sample containing the nanowire;
b) binding the probe DNA onto the quartz electrode;
c) performing hybridization of the linker DNA and the probe DNA by flowing the sample into a flow cell including the crystal electrode; And
d) analyzing that the resonance frequency of the quartz electrode is shifted as the weight of the quartz electrode is increased by the hybridization of step c)
Wherein the step a) is performed by mixing the linker DNA with a reducing agent to activate the thiol group of the linker DNA and then binding the activated thiol group to the silver nanowire. . delete The method according to claim 1,
Wherein the reducing agent is selected from the group consisting of tris (2-carboxyethyl) phosphine, dithiothreitol, dithiolbutylamine, beta-mercaptoethanol and mixtures thereof. The method according to claim 1,
Wherein the resonance frequency shift of step (d) is calculated by the following equation (1):
&Quot; (1) "

Where f ₀ is the natural frequency of the non-hybridized quartz electrode, A is the active area of the quartz electrode, and ρ _q and μ _q are the density and shear modulus of the quartz electrode, respectively. The method according to claim 1,
The binding of the probe DNA to the quartz crystal electrode in step b) may be confirmed by observing the surface of the quartz crystal electrode with an atomic force microscope or by calculating the surface roughness of the quartz crystal electrode surface in an image obtained by the atomic force microscope Wherein the method comprises the steps of: The method according to claim 1,
Wherein the binding between the silver nanowire and the linker DNA in step (a) is confirmed by analyzing a UV-Visible absorption spectrum of the silver nanowire. The method according to claim 1,
1 ng / ml. ≪ / RTI >

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