KR102190393B1 - Paper-based colorimtric sensor for on-site high efficient, rapid and visual detection of mercuric ions and on-site high efficient, rapid and visual detection of mercuric ions - Google Patents

Abstract

A paper-based colorimetric sensor kit is available for quick and easy visual field diagnosis of mercury. Colorimetric sensor kit is generated from a circular template for rolling circular amplification (RCA), a primer that does not hybridize to the template when mercury ions are hybridized with a primer that hybridizes to the template, DNA polymerase, and a circular template for the rolling circular amplification. It includes a sensing material kit and radial paper chromatography including nanoparticle probes labeled on the DNA coil.

Description

A paper-based colorimetric sensor kit for quick and simple visual spot diagnosis of mercury and a quick and easy visual spot detection method of mercury using it {PAPER-BASED COLORIMTRIC SENSOR FOR ON-SITE HIGH EFFICIENT, RAPID AND VISUAL DETECTION OF MERCURIC IONS AND ON-SITE HIGH EFFICIENT, RAPID AND VISUAL DETECTION OF MERCURIC IONS}

The present disclosure relates to a sensor for diagnosis of mercury ions in water and a detection method using the same, and more specifically, a paper-based colorimetric sensor kit for quick and simple visual field diagnosis of mercury ions in water, and rapid and simple pathogenic bacteria using the same It relates to the visual field detection method.

Mercury is one of the toxic heavy metals that pose a serious threat to the global environment and human health. In particular, mercury ions (Hg ²⁺ ) are one of the most important factors in water pollution. Mercury ions (Hg ²⁺ ) are converted to methyl mercury through microbial biosynthetic pathways, which are easily absorbed by bacteria, plankton, and fish. It causes serious damage to the human brain, nervous system, kidney and endocrine system.

Currently, the most common methods used for mercury detection include Inductively Coupled Plasma Mass Spectrometry (ICP-MS), Atomic Absorption Spectroscopy (AAS), and Gas Chromatography (GC). This method can detect a wide range of metal ions with high sensitivity and selectivity, but it is expensive and requires sophisticated equipment and skilled personnel. In addition, detection is time-consuming and requires labor-intensive procedures. Therefore, there is a need to develop a method that can be quickly and easily diagnosed on-site with the naked eye, and is simple and convenient, and can be detected with the naked eye immediately in the field.

It is intended to provide a paper-based colorimetric sensor kit for mercury detection that can quickly and simply detect mercury in real time in real time and does not require complex expensive measuring equipment, and a quick and simple visual field detection method of mercury using the same. .

In addition to the above problems, the embodiments according to the present invention may be used to achieve other tasks not specifically mentioned.

The paper-based colorimetric sensor kit for quick and simple visual field diagnosis for mercury detection according to the embodiments is a circular template for rolling circular amplification (RCA), a primer that hybridizes to the template, and does not hybridize to the template when mercury ions are combined. It includes a sensing material kit and radial paper chromatography including a primer, a DNA polymerase, and a nanoparticle probe labeled on a DNA coil generated from a circular template for rolling circular amplification.

The quick and simple visual field diagnosis method for mercury detection according to the embodiments is a primer that hybridizes to a circular template for rolling circular amplification. When mercury ions bind, a primer that does not hybridize to the template and a detection solution are mixed and reacted, and the mixing In the reaction solution, a circular template for rolling circular amplification, a DNA polymerase, and a nanoparticle probe labeled on a DNA coil generated in the circular template for rolling circular amplification, and dNTP are mixed to proceed with the RCA reaction, and the RCA reaction is completed. It includes dropping the solution on radial paper chromatography, drying, and observing the concentric circles generated by the paper chromatography.

The paper-based colorimetric sensor kit according to the present disclosure uses a handheld spectrophotometer to measure mercury ions (Hg ²⁺ ) without being affected by lighting conditions such as weather, sunlight intensity, and angle at the reading point through a very simple and robust measurement. The concentration can be easily measured. That is, when the paper-based colorimetric sensing method according to an embodiment of the present invention is used, the mercury ion (Hg ²⁺ ) concentration can be determined without requiring a bulky and expensive tool for reading, and is simple, portable, and cost-effective. Providing an approach allows quick and easy visual on-site diagnosis.

FIG. 1 is a conceptual diagram illustrating a method for detecting a simple visual field of mercury using a paper-based colorimetric sensor kit according to an embodiment of the present invention.
2 shows the results of PAGE analysis to determine whether a circular template was generated.
3 is a transmission electron microscopy (TEM) photograph when only the Au NP probe is present.
4 is a TEM photograph showing that the Au NP probe is aggregated on ss DNA.
5 shows the results of measuring the characteristics of the Au NP probe by dynamic light scattering.
6 shows the result of measuring the characteristics of the Au NP probe as a zeta potential.
7 shows the yield of the ssDNA coil according to the amount of the circular template-(T)12 primer complex.
8 shows a digital image of a concentric ring formed on a paper after reacting different metal ions with a primer.
9 shows a bar graph of color intensity (ΔE) quantified by a handheld spectrophotometer.
10 is a fluorescence intensity graph showing the results of measuring selectivity of different metal ions and primers.
11 is a digital image showing the colorimetric response on paper for different concentrations of mercury ions.
12 shows a graph quantifying colorimetric reactions according to eight different mercury ion concentrations using a spectrophotometer.
13 shows a graph in which colorimetric reactions according to five different concentrations of mercury ions are quantified using a spectrophotometer.
FIG. 14 is a digital image of measuring colorimetric reaction on paper after addition of mercury ions (Hg ²⁺ ) at different concentrations of 0, 50, 250, 500 and 1000 nM to tap water.
FIG. 15 shows a graph in which the colorimetric reaction on paper was quantified using a spectrophotometer after addition of mercury ions (Hg ²⁺ ) at different concentrations of 0, 50, 250, 500 and 1000 nM to tap water.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings so that those of ordinary skill in the art may easily implement the present invention. The present invention may be implemented in various different forms, and is not limited to the embodiments described herein.

When a certain part in the specification "includes" a certain element, it means that other elements may be further included unless otherwise specified. Since the size and thickness of each component shown in the drawings are arbitrarily shown for convenience of description, the present invention is not limited as shown.

FIG. 1 is a conceptual diagram illustrating a method for detecting a simple visual field of mercury using a paper-based colorimetric sensor kit according to an embodiment of the present invention.

The paper-based colorimetric sensor kit according to an embodiment of the present invention includes a sensing material kit for detecting mercury ions (Hg ²⁺ ) and a paper for colorimetric reaction.

The sensing material kit 10 includes a circular template 12 for rolling circle amplification (RCA), a primer 14, a DNA polymerase 16, and a nanoparticle probe 18.

First, in order to test whether mercury ions are present, the test object is reacted with the primer 14 (S1).

When the test object is in the same solution as tap water, the test object is mixed in the reactor containing the sensing material kit 10. If the test object is not a solution, the sampled test object is dissolved in distilled water and then mixed in the reactor contained in the sensing material kit 10.

The reaction between the primer 14 and the test object may be performed at room temperature for 10 to 30 minutes.

Then, the RCA reaction proceeds (S2). For the RCA reaction, the circular template 12, the DNA polymerase 160, the nanoparticle probe 18, and the dNTP were mixed in the reactor 20 containing the solution in which the test object and primer were mixed, and 30 minutes at 30°C. RCA reaction can be carried out by incubating for 60 minutes.

RCA is an isothermal enzymatic process mediated by a specific DNA polymerase in which long ssDNA molecules are synthesized from a short circular ssDNA template using one DNA primer. RCA is a powerful sensing system because it spatially condenses to create a single-stranded DNA coil of micron-sized DNA concatemers that can be detected as a single amplified molecule. In some common RCA detection sensors, the RCA response is induced by the connection of the lock probe, which depends on the presence of the object to be detected.

On the other hand, in an embodiment of the present invention, a selective reaction between mercury ions (Hg ²⁺ ) and primer 14 is used. The primer 14 hybridizes to the circular template 12 for RCA under normal conditions. However, when mercury ions (Hg ²⁺ ) are present, mercury ions (Hg ²⁺ ) bind to the primer 14 and inhibit the primer 14 from binding to the circular template 12.

The circular template 12 for RCA is a template in which 50 to 80 bp of single-stranded DNA (ss DNA) is formed in a circular shape.

In the absence of mercury ions (Hg ²⁺ ), the RCA reaction is induced by DNA polymerase (eg, Phi29 DNA Polmerase) 18 when the primer 14 in a steady state is hybridized to the circular template 12. During the RCA reaction, under isothermal conditions, a single-stranded DNA coil 19 made of micron-sized DNA concatemerics is produced. When the resulting single-stranded DNA coil 19 is produced, the probe 18 is labeled on the DNA coil 19.

On the other hand, when mercury ions (Hg ²⁺ ) are present, mercury ions (Hg ²⁺ ) are bound to the primer 14 to prevent hybridization of the primer 14 to the circular template 12. For example, the primer 14 is thymine (Thymine) oligonucleotides in the case of mercury ions to a combination (Hg ²⁺⁾ thymine -Hg ²⁺ - form a coordination bond of thymine, and since the changes in the mold This circular double stranded complex It becomes impossible to hybridize to (12).

The probe 18 may be formed of an oligonucleotide capable of hybridizing with the single-stranded DNA coil 19 and nanoparticles (NPs) exhibiting a predetermined color. For example, the NP may be Au NP, and the oligonucleotide may be functionalized to Au NP using streptavidin-biotin.

When the RCA-completed solution is dropped onto the radial paper chromatography 30 and dried, the color change on the paper can be quickly and simply measured using a handheld spectrophotometer within a few seconds (S3).

In the radial paper chromatography (30), the solution to be measured is dropped in the center in a radial form rather than a lateral form, and the presence and extent of mercury ions (Hg ²⁺ ) are easily determined using the principle that the fluidity is different depending on the molecular size and weight. I can confirm.

The resolution may vary depending on the material, pore size, hydrophilicity or hydrophobicity characteristics of the radial paper chromatography 30. Radial paper chromatography 30 may be paper chromatography made of PVDF (polyvinylidene fluoride) NC (nitrocellulose), or the like.

When the RCA reaction is completed solution is dropped onto the radial paper chromatography 30, the reaction solution moves from the center of the paper toward the periphery based on capillary action. The components of the solution are separated radially in the mobile phase, and spots of different compounds are formed as concentric rings in the stationary phase. In the detection system, the amount of the DNA coil and Au NP probe complex varies depending on the concentration of mercury ions (Hg ²⁺ ). When the concentration of mercury ions (Hg ²⁺ ) is large, the color of the concentric center spot 40 appears strong. On the other hand, when mercury ions (Hg ²⁺ ) are absent or the concentration is small, free Au NPs diffuse far in the radial direction, resulting in a strong color of the outer ring 50 of the concentric circle. Thus, a handheld spectrophotometer can be used to quantify the color intensity of each spot.

The following experimental examples and drawings are provided in order to better understand the conceptual aspects and methods of the embodiments of the present invention, and to elaborate operations and effects. However, these experimental examples are only presented as examples of the invention, and the scope of the invention is not limited thereby.

Experimental example

Manufacture of round molds

For the synthesis of circular template ssDNA, 100 μM of linear ssDNA (5′-phosphate GTCCTCAGTCCCAATAGAAGCGGAGCTTCAAAAAAAAAAAAACGTCTGAAGAGG) 5 μL of 100 μM of linear ssDNA having a length of 54 bp (100 μL DNA ligase) 4 μL, distilled water DDW) 71 μL, 1 mM ATP 5 μL, and 50 mM MnCl ₂ 5 μL) were linked at 60 for 8 hours and inactivated at 80 for 10 minutes. Then, in order to remove the remaining linear ssDNA, 6 μL of 20 U/μL exonuclease I and 3 μL of 10 U/μL exonuclease III were added, followed by incubation at 37 for 4 hours and at 80 for 20 minutes. The resulting product was raised and purified using an Oligo Clean-Up and Concentration Kit, and confirmed by 15% denatured urea-polyacrylamide gel electrophoresis (PAGE) for 75 minutes at 100V with 1xTBE buffer and then 1xSYBR Green Gel stained for 15 min with II. The results are illustrated in FIG. 2.

Preparation of Au NP probe

To prepare an Au NP probe, the surface of streptavidin-conjugated Au NP (SA-Au NP) was functionalized with biotinylated oligonucleotides. The shape of the Au NP probe was observed using a transmission electron microscope (TEM) at an acceleration voltage of 80 kV.

Specifically, the SA-Au NP solution (300 μL, 10OD) was mixed with 100 μM biotinylated ssDNA probe (5′-TTTTTTTTTTTT-C6SP-biotin). Then, the mixture was incubated with shaking at room temperature (18-23 ℃) for 3 hours and incubated at 4 ℃ overnight. To obtain a purified DNA probe modified with Au NP, the solution was washed 3 times through centrifugation at 6000 x g for 30 minutes, and resuspended in 1x PB buffer at pH 7.5. Two washing steps were performed using distilled water. The shape of the Au NP probe was examined by TEM.

As can be seen in Figure 3, the Au NP probe is well dispersed in the solution and is spherical. In contrast, as can be seen in FIG. 4, aggregation of the Au NP probe is observed during the hybridization reaction of the Au NP probe and ssDNA. It is interpreted that this is because the oligonucleotide of the Au NP probe binds complementarily with ssDNA.

5 and 6 show the results of measuring the characteristics of the Au NP probe by dynamic light scattering (DLS) and zeta potential. After introducing a negatively charged biotinylated oligonucleotide onto the surface of SA-Au NP, it can be seen that the size and surface charge of the Au probe were changed. As shown in FIG. 5, it can be seen that the average diameter of the particles increased from 33.21 nm to 35.29 nm, and as shown in FIG. 6, the negative charge increased from -24.8 mV to -33.8 mV. These results indicate that oligonucleotides have been successfully attached to the surface of SA-Au NPs and are more stabilized by non-specific aggregation due to the increased negative surface charge.

RCA reaction test using prototype template and (T)12 primer complex

An experiment was performed to confirm DNA amplification through the formation of a circular template-(T)12 primer complex. Various concentrations of the prototype template and the (T)12 primer complex were tested under the experimental conditions of RCA reaction, and SYBR Green II was used as a dyeing reagent to determine the amount of amplified ssDNA coil.

Specifically, the fluorescence intensity of SYBR Green II was measured after RCA reaction to confirm the amount of ssDNA coil according to the amount of the circular template-(T)12 complex. First 80 μL of total reaction mixture (10 × 8 μL of buffer, 7.5 μL of 10 MM dNTP, 25 U/μL) containing various concentrations (6, 3, 1.5, 0.75, 0.375 and 0 μM) of the prototype template-(T)12 complex. phi29 DNA polymerase 3 μL and DDW 41.5 μL) were reacted at 30° C. for 90 minutes and inactivated at 65° C. for 10 minutes. Next, 1 μL of 100 × SYBR Green II was added, and after incubation for 30 minutes at room temperature, fluorescence intensity was measured using a multimode microplate reader (SpectraMax i3x, Molecular Devices, Sunnyvale, CA, USA).

7 shows the yield of the ssDNA coil according to the amount of the circular template-(T)12 primer complex. As the amount of the circular template-(T)12 primer complex increased from 0 to 6 μM, the fluorescence intensity of SYBR Green II also increased. These results indicate that the circular oligonucleotide successfully acts as a template for RCA and that the amount of ssDNA coil produced by RCA is proportional to the amount of the circular template-(T)12 primer complex.

Selectivity to mercury in paper-based colorimetric sensing method

Mercury ions, three each test a variety of metal ions as well as ions of mercury (Hg ²⁺⁾ in order to determine the selectivity of the proposed analysis methods on (Hg ²⁺⁾ to drop the solution on the NC membrane dried colorimetric intensity of concentric rings Compared. The results are shown in FIG. 8. The final concentration of each metal ion was 1 μM.

Specifically, to evaluate the selectivity for mercury ions, 16 metal ions (Li ⁺ , Mn ²⁺ , Ag ⁺ , Pb ²⁺ , Co ²⁺ , Ba ²⁺ , Ca ²⁺ , K ⁺ , Ni ²⁺ , Cd ²⁺ , Zn ²⁺ , Fe ²⁺ , Mg ²⁺ , Na ⁺ , Cu ²⁺ and Hg ²⁺ ) were tested. First, 5 μL of a 200 nM (T)12 primer solution was mixed with 5 μL of 16 μM of each metal ion or 5 μL of a metal ion mixture, followed by incubation at room temperature for 30 minutes. Then, the RCA reaction mixture [150 nM circular template 10 μL, 10 x reaction buffer 8 μL, 10 mM dNTP 7 μL, 10 U/μL phi29 DNA polymerase 2.5 μL, DDW 12.5 μL and Au NP probe (3.3 OD)] was added. And incubated at 30° C. for 30 minutes and 65° C. for 10 minutes. Then, 10 μL of each reaction solution was dripped and dried on the NC membrane. The results were analyzed using a handheld spectrophotometer (RM200QC, X-Rite Co., Neu-Isenburg, Germany). The spectrophotometer uses CIE L * a * b * (CIELAB), which represents the color space specified by the Commission Internationale del'Eclairage (CIE). CIELAB describes all colors visible to the human eye and is provided as a device-independent model for reference. When expressing colors in CIELAB, L * defines brightness, a * stands for red/green values, and b * stands for yellow/blue values. The value of ΔE represents the total color difference calculated by the following equation.

ΔE = [(ΔL *) 2 + (Δa *) 2 + (Δb *) 2] 1/2

* Fig. 8 shows a digital image of a concentric ring formed on paper. 9 shows a bar graph of color intensity (ΔE) quantified by a handheld spectrophotometer. The color density of the concentric rings on the paper was easily distinguished by the naked eye and could be measured numerically using a handheld spectrophotometer. In addition, when measuring the color density of the concentric rings on the paper, it can be seen that since the handheld spectrophotometer completely contacts the paper, constant measurement is possible regardless of external lighting conditions. This feature represents an advantage as a sensor for field detection of mercury ions (Hg ²⁺ ).

Looking at Figure 8, the color depth of the inner concentric ring was lower only from the mixture of two samples containing the sample and the ionic mercury (Hg ²⁺⁾ contained only mercury ions (Hg ^2+). This indicates that RCA inhibition occurred due to mercury ions (Hg ²⁺ ). On the other hand, in the case of the sample containing mercury ions (Hg ²⁺ ), since Au NPs were not bound to the DNA coil, higher color intensity was exhibited at the edge of the concentric circle. This result demonstrates that the proposed analytical method is selective for mercury ions (Hg ²⁺ ).

Also, the selectivity of the (T)12 primer for mercury ions (Hg ²⁺ ) is shown in FIG. 10. The high fluorescence value of SYBR Green I was found only in samples containing only mercury ions (Hg ²⁺ ) and mixtures containing mercury ions (Hg ²⁺ ) when compared to the values of other metal ion samples of the same concentration. This indicates that the single-stranded primer was changed into a double-stranded complex due to the coordination of thymine-Hg ²⁺ -thymine. As a result, it can be seen that the (T)12 primer bound to the mercury ion (Hg ²⁺ ) was unable to bind to the circular template, thereby inhibiting the RCA reaction.

Mercury ions (Hg) using a paper-based colorimetric sensing method ²⁺ ) Of

Fig. 11 is a digital image after being dropped on NC paper and dried. Referring to FIG. 11, it can be seen that in the absence of mercury ions (Hg ²⁺ ), an intense red spot appears in the center of a concentric circle, and the intensity of the spot decreases as the concentration of mercury ions (Hg ²⁺ ) increases. have. In contrast, it can be seen that the color intensity of the edge ring of the concentric ring increases as the concentration of mercury ions (Hg ²⁺ ) increases.

The results of quantifying the red spot at the center of the concentric circle using a handheld spectrophotometer are shown in FIGS. 12 and 13. 12 shows measurement results of nine samples of various mercury ions (Hg ^{2 +} ) concentrations (0, 50, 250, 500, 1000, 1500, 2000, 2500 and 3000 nM). The color density (ΔE) of the spot at the center of the concentric circles on the paper was measured as 19.2, 18.6, 16.8, 13.8, 10.2, 7.3, 6.1, 5.3 and 4.4, respectively, and the ΔE value was determined as the concentration of mercury ions (Hg ²⁺ ) increased. Tended to decrease accordingly.

13 shows the results performed at a concentration of mercury ions (Hg ²⁺ ) in the range of 0 to 1000 nM. Six samples (0, 50, 250, 500, 1000 and 1500 nM) with different mercury ions (Hg ²⁺ ) concentrations were tested three times and the average value is shown. Referring to FIG. 13, the average of ΔE values was 18.1±0.05, 17.5±0, 16.1±0.25, 14.4±0.77, 11.2±0.86, and 8.2±0.57. A linear curve for the concentration range of 0 to 1500 nM mercury ions (Hg ²⁺ ) and ΔE values was obtained, and the detection limit of mercury ions (Hg ²⁺ ) at a signal-to-noise ratio of 3 was calculated as 21.8 nM (n = 3). The measurement time using a handheld spectrophotometer was sufficient in a few seconds. Therefore, it can be seen that the total analysis time including the RCA reaction and measurement is around 90 minutes, which is very quick and simple measurement.

Mercury ions in tap water (Hg ²⁺ ) Of detection

To demonstrate the applicability of this detection method in real samples, the concentration of mercury ions (Hg ²⁺ ) was added to tap water at different concentrations of 0, 50, 250, 500, 1000 and 1500 nM, and paper-based colorimetric sensing method was used. Three samples were tested for each concentration. Before mercury mixing, tap water was prepared by filtration through an NC syringe filter membrane having a pore size of 0.22 μm.

Colorimetric test results are shown in FIGS. 14 and 15. The averages of ΔE values measured for each concentration of mercury ions (Hg ²⁺ ) of 0, 50, 250, 500, 1000 and 1500 nM were 18.27±0.05, 17.33±0.83, 16.57±0.48, 14.5±0.22, 11.07±0.52, respectively. 8.77±0.68. A linear curve of ΔE values was obtained in the range of 0 to 1500 nM Hg ²⁺ , and the detection limit at signal-to-noise ratio 3 was calculated as 22.4 nM (n = 3).

Thus, using the paper-based colorimetric sensing method according to an embodiment of the present invention, mercury ions are not affected by lighting conditions such as weather, sunlight intensity, and angle at the reading point through very simple and robust measurements using a portable spectrophotometer. The concentration of (Hg ²⁺ ) can be easily measured. That is, when the paper-based colorimetric sensing method according to an embodiment of the present invention is used, the mercury ion (Hg ²⁺ ) concentration can be determined without requiring a bulky and expensive tool for reading, and is simple, portable, and cost-effective. Providing an approach allows quick and easy visual on-site diagnosis.

In the above, preferred embodiments of the present invention have been described, but the present invention is not limited thereto, and it is possible to implement various modifications within the scope of the claims, the detailed description of the invention, and the accompanying drawings. It is natural to fall within the scope of

10: sensing material kit 12: circular mold
14: primer 16: DNA polymerase
18: nanoparticle probe 19: single stranded DNA coil
30: radial paper chromatography

Claims (9)

delete delete delete delete A step of preparing a primer mixture by mixing a detection solution and a primer to prepare a primer mixture, wherein the primer is hybridized to a circular template for rolling circular amplification (RCA) when mercury ions do not exist, and when mercury ions are present, the primer and the mercury Preparing a primer mixture that is a primer that inhibits hybridization of the primer to the circular template by binding of ions;
A step of performing a rolling circular amplification reaction by mixing a circular template for rolling circular amplification (RCA), a DNA polymerase, and a nanoparticle probe and dNTP labeled on a DNA coil generated from the circular template for rolling circular amplification in the mixed solution ;
Dropping the rolling circular amplification reaction solution onto radial paper chromatography and then drying it; And,
Including the step of observing the concentric circles generated in the paper chromatography,
When mercury ions are present in the detection solution, the primer and the mercury ions are bound to prevent hybridization of the primer to the circular template, and the RCA reaction does not occur, so that the nanoparticle probe is present in a free state, and the nanoparticles in the free state. The particle probe diffuses far in the radial direction of the paper chromatography, so that the color of the outer ring of the concentric circle appears strong,
If mercury ions do not exist in the detection solution, the RCA reaction occurs and the nanoparticle probe is labeled on the generated single-stranded DNA coil, so the color of the concentric center spot appears strong,
Observing the concentric circles generated in the paper chromatography is quick and simple visual field detection of mercury ions, including calculating the concentration of the mercury ions by quantifying the color intensity of the red spot in the center of the concentric circles using a portable spectrophotometer Way. The method of claim 5,
The primer is a thymine oligonucleotide, which specifically binds to the mercury ions or, if there is no mercury ions, binds to the circular template. The method of claim 5,
The nanoparticle probe is a quick and simple visual field detection method of mercury ions, which are Au nanoparticles representing color. The method of claim 7,
The Au nanoparticles are functionalized SA-Au NPs with Au nanoparticles immobilized on the surface of streptavidin with biotinylated oligonucleotides, and a quick and simple visual field detection method of mercury ions capable of hybridizing with DNA coils. delete

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