KR20040043677A - Device and method for detecting a nucleic acid hybridization using hole-type nucleic acid chips - Google Patents

Abstract

PURPOSE: An apparatus and method for detecting nucleic acid hybridization using hole-type nucleic acid chips are provided, thereby increasing the amount of probe nucleic acids fixed by increasing the surface area of a nucleic acid chip because nucleic acids are fixed on the hole-type construct. CONSTITUTION: An apparatus for detecting nucleic acid hybridization using hole-type nucleic acid chips comprises (1) a hole-type nucleic acid chip where the probe nucleic acid is fixed; (2) a reaction vessel for efficiently supplying and reacting a probe nucleic acid solution, buffer solution, target nucleic acid containing unknown sample, intercalator solution and transition metal complex solution; (3) electrodes and an electric power supplying device; and (4) a light measuring device for detecting luminescence of the intercalator and transition metal complex. A method for detecting nucleic acid hybridization comprises the steps of: fixing a probe nucleic acid on the hole inner wall(16) of the nucleic acid chip; washing the hole inner wall(16) with phosphate buffer solution and distilled water and supplying a target nucleic acid containing sample to hybridize them; washing the hole inner wall(16) with buffer solution to remove an unhybridized sample, supplying intercalator solution to bind the intercalator with the hybridized double strand nucleic acid; washing the hole inner wall(16) with buffer solution to remove an unbound intercalator; and positioning the hole type nucleic acid apart from a distance of a working electrode(15), supplying transition metal complex solution, and applying electric voltage to the surface of the electrode(15) to induce oxidation-reduction between the intercalator and transition metal complex.

Description

Nucleic acid hybridization detector and nucleic acid hybridization detection method using hole-type nucleic acid chip {DEVICE AND METHOD FOR DETECTING A NUCLEIC ACID HYBRIDIZATION USING HOLE-TYPE NUCLEIC ACID CHIPS}

The present invention relates to a nucleic acid hybridization detection method using electrochemiluminescence, and more particularly, to using a hole-type nucleic acid chip, and more effectively electrochemically by immobilizing a probe nucleic acid on an inner wall of a hole and placing it around an indicator electrode. The present invention relates to a method of inducing a luminescent response and detecting the nucleic acid.

Recently, as a method for analyzing a nucleic acid having a specific base sequence, a nucleic acid chip in which a large amount of genetic information is arranged on a solid substrate has been developed. This is a high density arrangement of nucleic acids known to know the information about the base sequence on a narrow solid substrate, the nucleic acid bound or immobilized on the surface is called a probe nucleic acid, the unknown containing a nucleic acid having an unknown sequence The sample is reacted to determine whether hybridization with the probe nucleic acid is used to find out information about the nucleic acid in the unknown sample. At this time, the nucleic acid in the unknown sample that is hybridized with the probe nucleic acid to form a double helix form is called a target nucleic acid.

In order to know the result of hybridization of nucleic acids, a method of labeling a target nucleic acid using a radioisotope or a fluorescent dye is generally widely used. ³² P or ³⁵ S is used as the radioisotope, and the binding state is determined using a photographic plate. This method has a disadvantage in that it is difficult to secure safety of the radioisotope, low resolution, and long analysis time. Have. In addition, when fluorescent dyes are used, an expensive laser scanner for reading out specific wavelengths generated by exciting the fluorescent dyes is required.

In addition to the above two methods, there is a method for measuring hybridization of nucleic acid by an electrochemical method. This is to measure the electrochemical properties resulting from the redox reaction of the transition metal complex and nucleic acid with electrochemical activity (U.S. pat. No. 5312527). Such an electrochemical method is possible with a low cost measuring device of a simple system, but has a disadvantage of poor sensitivity.

The detection technique using electrochemiluminescence, which is a method used in the present invention, provides a redox reaction with an intercalator that specifically binds a hybridization reaction between a probe nucleic acid and a complementary nucleic acid to a transition metal complex and a nucleic acid having a double-helix structure. It is by measuring the light emission through. This method eliminates the need to covalently separate and purify the sample with the label in advance, and eliminates the need for external light sources such as lasers or lamps to detect the label. It becomes possible. However, when the nucleic acid is immobilized on the electrode to which the applied voltage is applied, it is difficult to apply a stable voltage, and there is a need to improve the process of processing a sample required to cause a hybridization reaction and light emission.

In addition, in the measurement of hybridization using electrochemiluminescence, when the probe nucleic acid is immobilized directly on the working electrode, damage to the probe nucleic acid occurs in the process of applying a constant voltage to the electrode, or the probe nucleic acid is interposed between transition metal complexes on the electrode surface. Inhibition of the oxidation-reduction reaction may result in a decrease in electrochemiluminescence and inferior reproducibility.

The present invention solves the above problems by using a hole-type nucleic acid chip, and reduces the time required for total hybridization detection and the amount of samples required, thereby providing an effective nucleic acid hybridization detector and nucleic acid hybridization with a simple and excellent measurement sensitivity. It is an object to provide a detection method.

In addition, an object of the present invention is to provide a nucleic acid hybridization detector and a nucleic acid hybridization detection method capable of more easily measuring electrochemical luminescence by separating the hole inner wall and the electrode to which the probe nucleic acid is immobilized at regular intervals.

1 illustrates a hole type nucleic acid chip for fixing a probe nucleic acid.

2 shows the principle of electrochemiluminescence by an intercalator on a chip.

Figure 3 shows a method for measuring electrochemiluminescence using a nucleic acid chip of the hole method.

Figure 4 is a schematic diagram showing the position of the hole-type nucleic acid chip and the reference electrode when measuring the hybridization of nucleic acid.

Figure 5 shows the electrochemiluminescence measurement results of the hole-type nucleic acid chip using daunorubicin (daunorubicin).

Figure 6 shows the electrochemiluminescence measurement results of the hole-type nucleic acid chip using doxorubicin (doxorubicin).

Figure 7 shows the electrochemiluminescence measurement results of the hole-type nucleic acid chip using DAPI.

8 shows a method for simultaneously processing a sample in a plurality of hole-type nucleic acid chips, it is possible to effectively process a variety of samples through the flow of the fluid through the hole.

9 is a graph showing the change in electrochemical light emission of Ru (bpy) ₃ ²⁺ with pH.

*** Description of the main parts of the drawing ***

1: inner wall of hole to which probe nucleic acid is fixed, 2: substrate

3: inner wall of hole with gold thin film treatment, 4: probe nucleic acid

5: target nucleic acid, 6. intercalator

7: ruthenium solution, a transition metal complex, 8: hole-type nucleic acid chip

9: working electrode, 10: counter electrode

11: reference electrode, 12: potentiostat

13: transparent window of the measuring vessel, 14: optical measuring device

15: indicator electrode, 16: inner wall of the hole to which the nucleic acid is attached

17: transparent window, 18: light measuring device

The present invention relates to a nucleic acid hybridization detector which is more effective in introducing an electrochemiluminescence method and measuring the nucleic acid hybridization, and a method for hybridization detection of nucleic acid using the same.

This will be described in more detail as follows.

First, the nucleic acid detector according to the present invention

1) a hole-type nucleic acid chip portion to which a probe nucleic acid is fixed,

2) a container capable of efficiently supplying and reacting a transition metal complex solution, a buffer solution, an unknown sample containing a target nucleic acid, an intercalator, and the like,

3) electrode part and electric supply part, and

And 4) an optical measuring device portion for measuring the luminescence reaction of the intercalator and the transition metal complex.

In this case, the probe nucleic acid immobilized on the hole-type nucleic acid chip is a single-stranded oligonucleotide having a sequence complementary to the target nucleic acid of the sample to be measured, and the terminal nucleic acid is preferably induced to include a thiol group or an amine group. .

The hole-type nucleic acid chip is a hole that penetrates the substrate thickness to the substrate, and the nucleic acid material having various base sequences is fixed to the inner wall of the hole at a high density. As a substrate, PCB, glass, silicon wafer or other plastic Synthetic materials may be used, and it is preferable to coat the inner wall of the pores with a chemical or streptoavidine with poly-L-lycine, aldehyde or amine groups, and to add thiol functional groups or amine functional groups. The probe nucleic acid at the 5 'end is immobilized. Such a hole-type nucleic acid chip is used to find out information about nucleic acid in an unknown sample by hybridizing with a target nucleic acid in an unknown sample having a base sequence complementary to a nucleic acid (probe nucleic acid) immobilized on the inner wall of the hole. .

The intercalator used in the detection method according to the present invention specifically relates to a double-stranded nucleic acid formed by a hybridization reaction between a probe nucleic acid immobilized on the hole-type nucleic acid chip and a target nucleic acid on a sample solution. It is characterized in that the electrochemical luminescence is induced by inducing the oxidation-reduction reaction of the transition metal complex. The material used as the nucleic acid intercalator may bind specifically to the nucleic acid double helix structure and induce an oxidation-reduction reaction of the transition metal complex. As a material used as the intercalator, doxorubicin and daunorubicin are used. daunorubicin, nogalamycin, and DAPI (4 ', 6-Diamidino-2-phenylindole). The transition metal complexes include tris (2,2'-bipyridyl) ruthenium (II) [tris (2,2'-bipyridyl) ruthenium (II), Ru (bpy) ₃ ²⁺ ], tris (1,10) -Phenanthroline) ruthenium (II) ruthenium derivatives such as tris (1,10-phenanthroline) ruthenium (II), Ru (phe) ₃ ²⁺ ] or tris (2,2'-bipyridyl) osmium ( II) [tris (2,2'-bipyridyl) osmium (II), Os (bpy) ₃ ²⁺ ] and the like can be used.

The electrode portion may be composed of a three-electrode system or a two-electrode system. In the case of adopting a three-electrode system, the electrode portion is composed of a working electrode, a reference electrode, and a counter electrode that maintain a constant distance from the center of the inner wall of the hole where the nucleic acid is fixed. Gold electrodes are used as electrodes, Ag / AgCl electrodes or silver wires as reference electrodes, and platinum wires are used as counter electrodes. In the case of using the two-electrode system, the electrode portion is composed of an indicator electrode and a counter electrode having a constant distance from the center of the inner wall of the hole in which the nucleic acid is fixed, and the indicator electrode is the same as that of the three electrode, ground) or platinum wire is used. As the electricity supply device, a potentiostat for controlling an applied voltage between the reference electrode and the indicator electrode may be used.

In the fixing of the electrode and the nucleic acid chip, the center of the hole of the hole-type nucleic acid chip in which the circular indicator electrode and the actual probe nucleic acid are fixed is aligned so that the voltage application portions of the probe nucleic acid and the indicator electrode are positioned at regular intervals. This is because the intercalator bound to the double-helix type nucleic acid gives the proper distance to activate the electrochemical reaction of the transition metal complex, and the light scattered in all directions when the hole structure detects the electrochemical emission It is also intended to serve as a gathering tool. In addition, as described above, since the probe nucleic acid is fixed and the electrode portion are separated, various problems that occur when the probe nucleic acid is fixed directly on the working electrode, that is, the probe in the process of applying a constant voltage to the electrode The nucleic acid may be damaged or the oxidation-reduction reaction between the transition metal complexes on the electrode surface may be inhibited by the probe nucleic acid, thereby reducing electrochemical emission and reducing reproducibility.

In the case of using the hole-type nucleic acid chip as in the present invention, in the hybridization reaction, the intercalator treatment and the washing process, there is an advantage that several can be simultaneously performed using a small amount of solution. In other words, by placing the hole-type nucleic acid chips in a row to match the positions of the holes, connecting them to the channel, and then flowing the necessary solution, space and time can be saved and the experiment can be effectively performed with a small amount of samples.

In addition, the measuring method according to the present invention,

Arrange several hole-type nucleic acid chips in a row, set up holes to fit channels, and probe nucleic acid and buffer solutions through the channels. While supplying the sample containing the target nucleic acid, and the intercalator solution through a precision pump,

Immobilizing the probe nucleic acid on the inner wall of the hole of the nucleic acid chip;

Washing the inner wall of the hole of the nucleic acid chip in which the probe nucleic acid has risen to a monolayer with a phosphate buffer solution and distilled water, and then feeding and hybridizing a sample containing the target nucleic acid;

Washing with buffer solution to remove unhybridized sample, and then supplying an intercalator solution to bind an intercalator to said hybridized double helix nucleic acid;

Washing with buffer solution to remove unbound intercalator; And

A hole type nucleic acid chip comprising the hybridized double-stranded nucleic acid and an intercalator coupled thereto is positioned at a predetermined distance from the indicator electrode, and then supplied with a transition metal complex solution and applied an applied voltage on the electrode, The intercalator includes a step of inducing an oxidation-reduction reaction of the transition metal complex to cause electrochemical light emission. At this time, the temperature may be about room temperature. In addition, the amount of electrochemiluminescence tends to increase with increasing pH, as can be seen in Figure 9, it is most preferably performed between about pH 7 to 9.

At this time, the electrochemiluminescence is detected by a photodetector, and the detected light is transmitted to the PC to analyze the data as the light quantity change with time. Phosphate buffer (phosphate buffer saline), 5X SSC buffer or 1X SSC buffer solution may be used as the buffer solution. The electrochemical device may be configured by using a three-electrode system of a microelectrode of a fine patterned gold electrode, a counter electrode of platinum (Pt) wire, and a reference electrode of Ag / AgCl, or a microelectrode of gold electrode It is possible to construct and use a two-electrode system of a counter electrode made of ground or platinum wire. A device that applies a voltage between the reference electrode and the indicator electrode applies a voltage of +1.19 V to measure the light generated by the redox reaction between the ruthenium derivative and the intercalator. Photo-counter detector is used in the dark box to convert the change of light represented by the transition metal into a digital signal, displaying the amount of light over time on the PC and storing the data. It plays a role.

Nucleic acid (nucleic acid) to be detected in the present invention refers to oligonucleotide, DNA, RNA, PNA, cDNA and the like generically.

The present invention relates to a method for detecting nucleic acid hybridization in a hole-type nucleic acid chip using the above-described intercalator, and in particular, does not require any special pre-labeling operation on a target nucleic acid, and is fast, inexpensive and highly sensitive. Provide a method.

1 shows an example of a hole type nucleic acid chip. The hole inner wall 1 is treated with a thin film of gold. The material of the board | substrate 2 used the PCB 2.

2 shows the principle of electrochemiluminescence by an intercalator. A self-assembled monolayer is made by flowing a probe nucleic acid 4 having a thiol functional group at a 5'-phosphate position into the gold thin film-treated hole inner wall 3 of the hole-type nucleic acid chip of FIG. 1; SAM). As described above, when an unknown sample including the target nucleic acid (5) is administered to the hole nucleic acid chip in which the probe nucleic acid is immobilized on the inner wall of the hole, the target nucleic acid having a sequence complementary to the probe nucleic acid hybridizes with the probe nucleic acid, and the non-complementary nucleic acid. Are removed by a washing process. Here, the intercalator 6 is administered, and the intercalator mainly binds between the double-helix minor groove and major groove base pairs. Intercalators include doxorubicin, daunorubicin hydrochloride, nogalamycin, and DAPI (4 ', 6-Diamidino-2-phenylindole). The ruthenium solution (7), which is a transition metal complex, is flowed to the hole-type nucleic acid chip in which the intercalator is coupled, and an applied voltage of about 1.2 volts is applied to Ru (bpy) ₃ ²⁺ in the solution to oxidize Ru (bpy). ) ₃ ³⁺ derivative, which becomes an excited Ru (bpy) ₃ ^{2 + *} derivative through redox reaction with an intercalator, which returns to the grounded Ru (bpy) ₃ ²⁺ . When about 620 nm light is generated. Light generated at this time is detected by a photo detector.

FIG. 3 shows a nucleic acid detector including a hole-type nucleic acid chip and a method of measuring hybridization of nucleic acids by measuring electrochemical luminescence using the same. A nucleic acid chip 8 having a probe nucleic acid and a target nucleic acid fixed to an inner wall of the hole and having an intercalator coupled thereto is arranged in parallel with a working electrode 9. At this time, the indicator electrode uses a fine patterned circular gold electrode having a diameter smaller than the hole diameter of the nucleic acid chip, and the portions other than the circle are coated with an insulating film. A platinum wire may be used as the counter electrode 10, Ag / AgCl may be used as the reference electrode 11, and a potentiostat 12 may be used to apply a voltage. The window 13 of the measuring vessel is made of PDMS having excellent light transmission, injecting a ruthenium solution therein, and applying an applied voltage of +1.19 V to the electrochemical luminescence reaction to emit light of about 620 nm, This is measured through the optical measuring device 14. As an optical measuring device, a photodetector having an amplifying capability such as a photomultiplier tube (PMT) or an avalanche photodiode can be used, and a cooled CCD camera or the like can also be used. It is possible.

4 is a view showing the position of the hole-type nucleic acid chip and the indicator electrode in the measurement, the left side of the indicator electrode and the nucleic acid chip in the direction of the measuring device and the right side is viewed from above, the substrate and the nucleic acid patterned indicator electrode It shows the position of the fixed nucleic acid chip, the transparent window and the optical measuring device. A circular gold electrode is patterned on the substrate so that the center of the nucleic acid chip coincides with the center of the hole and used as the indicator electrode, so that the position of the indicator electrode and the nucleic acid chip is always constant. In this way, if the indicator electrode is immobilized, only the nucleic acid chip can be used for measurement. Nucleic acid materials immobilized on each chip are spaced adjacent to each other without being directly in contact with a voltage-applying indicator electrode, thereby providing stable results during electrochemical light emission. In addition, since light emitted from the electrochemical emission of ruthenium is reflected from the gold surface of the indicator electrode and the inner wall of the hole of the nucleic acid chip, the light is collected toward the measurement device, thereby increasing the amount of light that can be measured.

5, 6 and 7 show the results of experiments for detecting the hybridization results of nucleic acids through a nucleic acid detector including the hole-type nucleic acid chip. These experiments were performed using daunorubicin (FIG. 5), doxorubicin (FIG. 6), and DAPI (FIG. 7) as an intercalator. After applying an applied voltage of + 1.19V, relative light emission with time (t) was measured. (RLU) is measured. FIG. 5 shows the amount of luminescence obtained by applying a voltage of +1.19 V in 5 mM ruthenium solution after treatment with 1 mM Daunorubicin for 30 minutes to the hybridized nucleic acid chip, followed by washing with buffer and distilled water. . Electrochemical luminescence which appeared before and after the treatment of the intercalator of untreated probe nucleic acid, fixed single-stranded nucleic acid and fixed double-stranded nucleic acid was shown. As shown in the graph, it can be seen that the increase in the amount of light emitted after treatment with the intercalator and daunorubicin in the portion where the double-stranded nucleic acid is fixed is more pronounced than in the portion where the single-stranded nucleic acid is fixed. FIG. 6 shows the treatment of 1 mM doxorubicin, and FIG. 7 shows the result of the treatment of 1 mM DAPI. As in the case of FIG. 5, the light emission amount of the double-stranded nucleic acid portion was different from that of the single-stranded nucleic acid portion, and in particular, the DAPI showed a large difference under the same conditions.

FIG. 8 shows that the unknown sample, the washing buffer, and the intercalator solution flow through the hole to connect the solution inlet and outlet to the hole by arranging the hole type nucleic acid chips in a row. In the case of using a plate-shaped chip, a large amount of sample is used in the hybridization or washing step, and a lot of space is required to process several chips simultaneously. However, the hole-type nucleic acid chip can be used to process several pieces at the same time and a small amount of samples in a narrow space by lining up several chips, thereby reducing the time and cost used in each step. Will be.

The nucleic acid detector according to the present invention or a nucleic acid detection method using the same does not raise the nucleic acid on a working electrode or on a flat surface, and fixes the nucleic acid rather than the planar state of the chip by fixing the hole in a hole-shaped structure. Thus, the amount of probe nucleic acid that can be immobilized can be increased, thereby increasing the detection limit. In addition, since the fixing surface of the nucleic acid can be positioned at a predetermined distance from the working electrode, the reproducibility of the electrochemical luminescence experiment can be improved. In addition, since the hole-shaped chip collects light scattered in all directions toward the detection device, light quantity detection sensitivity is improved. In addition, by arranging the chips in the form of holes side by side, the flow of fluid through the holes is advantageous, so that the processes such as hybridization, washing, and intercalation can be performed very quickly and effectively.

Claims (18)

1) a hole-type nucleic acid chip portion to which a probe nucleic acid is fixed, 2) a reaction vessel capable of efficiently supplying and reacting probe nucleic acid solution, buffer solution, unknown sample containing target nucleic acid, intercalator solution and transition metal complex solution, 3) electrode part and electric supply part, and 4) A nucleic acid hybridization detector comprising a portion of an optical measuring device for measuring a luminescence reaction of an intercalator and a transition metal complex, wherein a portion of a hole-type nucleic acid chip to which a probe nucleic acid is fixed and an electrode portion are separately located. The nucleic acid hybridization detector of claim 1, wherein the 5 ′ terminal portion of the probe nucleic acid comprises a thiol functional group or an amine functional group. The method of claim 1, wherein the hole-type nucleic acid chip is drilled through the thickness of the substrate in the substrate of the PCB, glass, silicon wafer or other plastic composite material, the inner wall of the hole is gold or poly- A nucleic acid hybridization detector produced by applying a chemical product having L-lysine (poly-L-lycine), an aldehyde or an amine group or streptoavidine. The method according to claim 1, wherein the intercalator specifically binds to the nucleic acid double helix structure and can induce an oxidation-reduction reaction of the transition metal complex, doxorubicin, daunorubicin, or nogala. A nucleic acid hybridization detector selected from the group consisting of mycin (nogalamycin) and DAPI (4 ', 6-Diamidino-2-phenylindole). The method of claim 1, wherein the transition metal complex is tris (2,2'-bipyridyl) ruthenium (II) [tris (2,2'-bipyridyl) ruthenium (II), Ru (bpy) ₃ ²⁺ ] and Tris (1,10-phenanthroline) ruthenium (II) Ruthenium derivative or tris (2,2'- selected from tris (1,10-phenanthroline) ruthenium (II), Ru (phe) ₃ ²⁺ ] A nucleic acid hybridization detector wherein bipyridyl) osmium (II) [tris (2,2'-bipyridyl) osmium (II), Os (bpy) ₃ ²⁺ ]. According to claim 1, wherein the electrode portion is composed of a three-electrode system of a working electrode, a reference electrode (reference electrode) and a counter electrode at a constant distance from the center of the inner wall of the hole in which the nucleic acid is fixed A nucleic acid hybridization detector comprising a gold electrode as an indicator electrode, an Ag / AgCl electrode or silver wire as a reference electrode, and a platinum wire as a counter electrode. The method of claim 1, wherein the electrode portion is composed of a two-electrode of the indicator electrode and the counter electrode at a constant distance from the center of the inner wall of the hole in which the nucleic acid is fixed, a gold electrode as the indicator electrode, ground (ground) or A nucleic acid hybridization detector constructed using platinum wires. The nucleic acid hybridization detector of claim 1, wherein the electricity supply device is a potentiostat for controlling an applied voltage between the reference electrode and the indicator electrode. The nucleic acid hybridization detector of claim 1, further comprising a photomultiplier tube (PMT), an avalanche photodiode, or a cooled CCD camera. Using a nucleic acid hybridization detector comprising a hole-type nucleic acid chip portion according to claim 1, a reaction vessel, an electrode portion, an electricity supply portion, and an optical measuring device portion, several hole-type nucleic acid chips are arranged in a row and Center the channel and install the probe nucleic acid, buffer solution through that channel. While supplying the sample containing the target nucleic acid, the intercalator solution, the transition metal complex solution through the precision pump, Immobilizing the probe nucleic acid on the inner wall of the hole of the nucleic acid chip; Washing the inner wall of the hole of the nucleic acid chip in which the probe nucleic acid has risen to a monolayer with a phosphate buffer solution and distilled water, and then feeding and hybridizing a sample containing the target nucleic acid; Washing with buffer solution to remove unhybridized sample, and then supplying an intercalator solution to bind an intercalator to said hybridized double helix nucleic acid; Washing with buffer solution to remove unbound intercalator; And A hole-type nucleic acid chip including the hybridized double-stranded nucleic acid and an intercalator coupled thereto is positioned at a predetermined distance from the indicator electrode, and then supplied with a transition metal complex solution and applied an applied voltage on the electrode. And a nucleic acid hybridization detection method, wherein the intercalator induces an oxidation-reduction reaction of a transition metal complex to cause electrochemical emission. The nucleic acid hybridization detection method according to claim 10, wherein the probe nucleic acid uses an oligonucleotide having a 5 'terminal portion containing a thiol functional group or an amine functional group. The hole type nucleic acid chip according to claim 10, wherein the hole-type nucleic acid chip is made of a substrate made of PCB, glass, silicon wafer, or other plastic composite material, through which a hole penetrates the thickness of the substrate, and gold is formed on the inner wall of the hole. A nucleic acid hybridization detection method using a L-lysine (poly-L-lycine), an aldehyde or a chemical product having an amine group, or one prepared with streptoavidine. The electrode according to claim 10, wherein the electrode portion comprises a finely patterned three-electrode system of a reference electrode made of a gold electrode, an Ag / AgCl electrode, or a reference electrode made of silver wire and a counter electrode made of platinum wire. And a nucleic acid hybridization detection method, wherein the electrodes are arranged at regular intervals at the center of the inner wall of the hole in which the nucleic acid is fixed. 11. The method of claim 10, wherein the electrode portion is composed of a two-electrode system of an indicator electrode made of a gold electrode and a counter electrode made of a ground or platinum wire, and the electrodes are held at a constant distance from the center of the inner wall of the hole in which the nucleic acid is fixed. Nucleic acid hybridization detection method. The nucleic acid hybridization detection method according to claim 10, wherein a potentiostat that adjusts an applied voltage between a reference electrode and an indicator electrode is used as the electricity supply device. The nucleic acid hybridization detection method according to claim 10, wherein a phosphate buffer, 5X SSC buffer, or 1X SSC buffer solution is used as the buffer solution. The method of claim 10, wherein the intercalator, which specifically binds to the nucleic acid double helix structure and can induce an oxidation-reduction reaction of the transition metal complex, doxorubicin, daunorubicin, A nucleic acid hybridization detection method selected from the group consisting of nogalamycin and DAPI (4 ', 6-Diamidino-2-phenylindole). The method of claim 10, wherein as the transition metal complex, tris (2,2'-bipyridyl) ruthenium (II) [tris (2,2'-bipyridyl) ruthenium (II), Ru (bpy) ₃ ²⁺ ] and Tris (1,10-phenanthroline) ruthenium (II) Ruthenium derivative or tris (2,2'- selected from tris (1,10-phenanthroline) ruthenium (II), Ru (phe) ₃ ²⁺ ] A nucleic acid hybridization detection method using bipyridyl) osmium (II) [tris (2,2'-bipyridyl) osmium (II), Os (bpy) ₃ ²⁺ ].