Electrochemical biomolecular sensor based on DNA molecular clock and construction method thereof

Abstract

The application discloses a kind of electrochemical biomolecular sensor and construction method based on DNA molecular pendulum, including buffer solution, electrode, DNA double-stranded pendulum rod, the electrode, DNA double-stranded pendulum rod is set in buffer solution;Step voltage reduction signal molecule is inserted on the phosphate skeleton of the DNA double-stranded pendulum rod;One end of the DNA double-stranded pendulum rod is connected on electrode, and one single strand on the other end of the DNA double-stranded pendulum rod is connected with target aptamer.The application has wider detection range, higher sensitivity and lower detection limit, and has important significance for detecting protein, virus, nucleic acid and some small molecule substances.

Description

Technical Field

[0001] This invention relates to a method for preparing an electrochemical biomolecular sensor and its application, belonging to the field of electrochemical biosensor technology. Background Technology

[0002] Disease biomarkers (also known as biomarkers or markers) are biomolecules that indicate the presence or state of a specific disease or cellular changes. These biomarkers can be proteins (antigens, antibodies), hormones, enzymes, specific cell types, or other molecules, and can be detected in bodily fluids or tissues such as blood, urine, and tissue samples. Disease biomarkers have a wide range of applications, including early diagnosis, disease monitoring, treatment evaluation, and prognosis. With advancements in medical research, more and more disease biomarkers are being discovered and applied in clinical practice. However, some instruments used for these biomarkers are expensive and complex to operate. Therefore, developing an economical, practical, highly sensitive detection method with a low detection limit is crucial.

[0003] Electrochemical biosensors have experienced rapid development in recent years. They are highly sensitive, capable of detecting very low concentrations of analytes, and offer rapid response, providing measurement results quickly. They are also highly selective and portable, as electrochemical sensors can be designed to be small and lightweight, suitable for field detection and portable applications. Furthermore, they are simple to operate, and their manufacturing and operating costs are generally lower compared to other sensing technologies. These advantages have led to the widespread application of electrochemical sensors in various fields, including environmental monitoring, food safety, and medical diagnostics. Summary of the Invention

[0004] Purpose of the invention: To address the problems of complex and costly biomarker detection operations in current methods, this invention provides an electrochemical biomolecular sensor based on a DNA molecular pendulum.

[0005] Technical solution: To achieve the above objectives, the technical solution adopted by this invention is as follows:

[0006] An electrochemical biomolecular sensor based on a DNA molecular pendulum includes a buffer solution, an electrode, and a DNA double-stranded pendulum, wherein the electrode and the DNA double-stranded pendulum are disposed in the buffer solution. A step voltage reduction signal molecule is inserted into the phosphate backbone of the DNA double-stranded pendulum. One end of the DNA double-stranded pendulum is connected to the electrode, and a target aptamer is attached to one of the single strands at the other end of the DNA double-stranded pendulum.

[0007] Preferably, the step voltage reduction signal molecule is [Ru(NH3)6]. 3+ Signal reduction molecules.

[0008] Preferably, the electrode is a gold electrode, a silver electrode, a silver chloride electrode, or a platinum electrode.

[0009] Preferably, the buffer solution is one or a mixture of two or more of Tris-HCl solution or phosphate-buffered saline (PBS).

[0010] Preferably, the target aptamer is CEA, PSA, RNA, or ATP with a target.

[0011] Another objective of this invention is to provide a detection method for an electrochemical biomolecular sensor based on a DNA molecular pendulum. In chronocurrent (CA) mode, a DNA double-stranded pendulum swings towards an electrode under the influence of an applied step negative electric field. A step voltage reduction signal molecule is reduced under this voltage, generating a detectable Faraday current. When the target aptamer on the DNA double-stranded pendulum binds to the target marker, its hydrodynamic diameter increases, the swing time becomes longer, and the current decreases. By detecting changes in current, it can be determined whether a target marker has bound to the target aptamer on the DNA double-stranded pendulum, thereby achieving the detection of the target marker.

[0012] Another object of the present invention is to provide a method for constructing an electrochemical biomolecular sensor based on a DNA molecular pendulum, comprising the following steps:

[0013] Step S1: The electrode is ultrasonically treated in ultrapure water, then polished, and finally rinsed with ultrapure water.

[0014] Step S2 involves chemically cleaning and activating the polished electrode.

[0015] Step S3: Incubate the DNA double-stranded lever containing the target aptamer on the cleaned and activated electrode, remove the electrode and blow it clean, then use [Ru(NH3)6] to clean it. 3+ Signal-reducing molecules are incubated with the electrode to modify the DNA molecule. After removing the electrode and cleaning it, MCH is used to incubate the electrode to shield non-specific binding sites.

[0016] Step S4: After the MCH incubation electrode has been shielded from non-specific binding sites, rinse it with buffer solution and store it in buffer solution to obtain an electrochemical biomolecular sensor based on a DNA molecular pendulum.

[0017] Preferred method: In step S2, the polished and cleaned electrode is activated in H2SO4 using cyclic voltammetry, with continuous CV scanning at 0.03-0.7V / s, and the potential range is -0.2 to 1.6V.

[0018] Preferred: In step S3, 4.0-6.0 μL of 8-12 μM DNA double-stranded pendulum with target aptamer is incubated on the cleaned and activated electrode, and then placed in a -15-25℃ refrigerator for 0.5-1.5 h.

[0019] Preferred: In step S3, [Ru(NH3)6] is used. 3+ Signal reduction molecules were incubated with electrodes to modify DNA molecules and placed at -5 to -3°C for 0.5–1.5 h. Non-specific binding sites were shielded using MCH incubation electrodes and placed at -5 to -3°C for 0.5–1.5 h.

[0020] Compared with the prior art, the present invention has the following advantages:

[0021] (1) The inventors of this invention incubated double-stranded DNA with aptamers onto a gold electrode, and ruthenium redox signaling molecules ([Ru(NH4)]6) were used. 3+ As a modifying material, it is used in the fabrication of electrochemical biomolecular sensors. Based on the specific recognition of aptamers and target substances, the molecular arrangement increases in hydrodynamic diameter after binding to the target.

[0022] The longer time required for DNA molecules to oscillate after an electric field is applied results in a regular electrical signal that is correlated with the target concentration for detection. The sensor is simple to construct, sensitive, and reproducible.

[0023] (2) The electrochemical biomolecular sensor prepared by the present invention is used for the detection of disease biomarkers CEA, PSA, RNA and ATP. The sensor has high stability, good reproducibility, high sensitivity and wide linear range, and can realize rapid, highly sensitive and reproducible continuous detection. Attached Figure Description

[0024] Figure 1 A schematic diagram of the detection of an electrochemical biomolecular sensor for a DNA molecular pendulum.

[0025] Figure 2 The electrochemical DNA molecular pendulum sensing platform is used to detect disease biomarkers at different concentrations. (a) CEA, (b) mRNA21, (c) ATP. Detailed Implementation

[0026] The present invention will be further illustrated below with reference to the accompanying drawings and specific embodiments. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the invention. After reading this invention, any modifications of the invention in various equivalent forms by those skilled in the art will fall within the scope defined by the appended claims.

[0027] Example 1

[0028] An electrochemical biomolecular sensor based on a DNA molecular pendulum can be used for the detection of disease biomarker proteins, or other biomarker proteins, such as... Figure 1As shown, the device includes a buffer solution, an electrode, and a DNA double-stranded lever. In this embodiment, the electrode is a gold electrode. In another embodiment, the electrode is a silver electrode; in yet another, it is a silver chloride electrode; and in still another, it is a platinum electrode. The electrode and the DNA double-stranded lever are placed in a buffer solution, which is a Tris-HCl solution. A step voltage reduction signal molecule is inserted into the phosphate backbone of the DNA double-stranded lever. In this embodiment, the step voltage reduction signal molecule is [Ru(NH3)6]. 3+ In another embodiment, the step voltage reduction signal molecule is ruthenium hexaammonium chloride ([Ru(NH3)6]). 3 Cl3) signal reduction molecules. One end of the DNA double-stranded lever is connected to an electrode, and a target aptamer is attached to one of the single strands at the other end of the DNA double-stranded lever.

[0029] We utilize double-stranded DNA with aptamers as the working basis for the molecular pendulum, and the ruthenium redox signaling molecule ([Ru(NH3)6)) 3+ The aptamer can electrostatically interact with the phosphate backbone of double-stranded DNA, inserting itself into the DNA backbone and giving the DNA a positive charge. In chronoamperometry (CA) mode, under the influence of an applied step negative electric field (-0.5–0 V), the DNA oscillates towards the Au electrode. The signal molecule is then reduced under a step voltage, generating a detectable Faraday current. When the aptamer on the DNA molecule binds to the target marker, its hydrodynamic diameter increases, and the oscillation time becomes longer. According to the Cotterell equation, the longer the time, the smaller the current. Based on this principle, the detection of disease biomarkers can be achieved. Test results show that this electrochemical sensor has high sensitivity, low detection limit, and good stability.

[0030] This embodiment is based on a DNA double-stranded aptamer, where one single-stranded elongated target aptamer can recognize specific biomolecules. In chronoamperometry mode, the electron transfer process can be precisely recorded, thereby achieving the detection of the target substance. Electrochemical MS sensing detects bio-recognition events based on active, strong electric field-mediated electrical signal transmission. The sensor's signal molecule is based on [Ru(NH3)6]. 3+ To [Ru(NH3)6] 2+ This is achieved through reduction; therefore, the sensor's detection capability can be restored through reverse oxidation or slow natural oxidation (sensor regeneration). Compared to sensors that perform single-use detection, this allows for continuous multiple detections, resulting in longer operating times and making it more suitable for long-term detection, significantly improving sensor efficiency.

[0031] Existing MS sensors (oxidizing signal molecules) driven by positive electric fields are susceptible to interference from reducing substances in the biological environment (e.g., glutathione, ascorbic acid, uric acid, dopamine, hydrogen peroxide, etc.). This embodiment, however, uses [Ru(NH3)6]. 3+ DNA is modified, [Ru(NH3)6] 3+ Insertion into the phosphate backbone of DNA via electrostatic interactions allows for the regulation of the charge and molecular position on the DNA by controlling the modified strand and the concentration of the modifying molecule. The CA method was used to [Ru(NH3)6]... 3+ Reducing to [Ru(NH3)6] 2+ Compared with the CA method using positive potential, it can avoid interference from reducing substances, has better anti-interference ability, makes experimental results more accurate, and is more suitable for the detection and analysis of biomolecules in body fluids.

[0032] Example 2

[0033] This embodiment provides a detection method for an electrochemical biomolecular sensor based on a DNA molecular pendulum. In chronocurrent CA mode, under the action of an applied step negative electric field, the DNA double-stranded pendulum swings towards the electrode. The step voltage reduces the signal molecule, generating a detectable Faraday current. When the target aptamer on the DNA double-stranded pendulum binds to the target marker, its hydrodynamic diameter increases, the time required for oscillation increases, and the current decreases. By detecting the change in current, it can be determined whether the target marker has bound to the target aptamer on the DNA double-stranded pendulum, thereby realizing the detection of the target marker. If the target marker has bound to the target aptamer on the DNA double-stranded pendulum, the current gradually decreases. Therefore, if the current gradually decreases, it indicates that the target marker has bound to the target aptamer on the DNA double-stranded pendulum, indicating that the target marker has been detected.

[0034] Example 3

[0035] This embodiment provides a method for constructing an electrochemical biomolecular sensor based on a DNA molecular pendulum, including the following steps:

[0036] Step S1: The electrode is ultrasonically treated in ultrapure water, then polished, and finally rinsed with ultrapure water.

[0037] Step S2 involves chemically cleaning and activating the polished electrode.

[0038] In another embodiment, the polished and cleaned electrode was activated in H2SO4 using cyclic voltammetry, with continuous CV scanning at 0.03-0.7V / s, and the potential range was -0.2 to 1.6V.

[0039] Step S3: Incubate the DNA double-stranded lever containing the target aptamer on the cleaned and activated electrode, remove the electrode and blow it clean, then use [Ru(NH3)6] to clean it. 3+ Signal-reducing molecules are incubated with the electrode to modify the DNA molecule. After removing the electrode and cleaning it, MCH is used to incubate the electrode to shield non-specific binding sites.

[0040] In another embodiment, 4.0-6.0 μL of 8-12 μM DNA double-stranded pendulum with target aptamers was incubated on the cleaned and activated electrode, and then placed in a freezer at -15-25°C for 0.5-1.5 h.

[0041] In another embodiment, [Ru(NH3)6] is used. 3+ Signal reduction molecules were incubated with electrodes to modify DNA molecules and placed at -5 to -3°C for 0.5–1.5 h. Non-specific binding sites were shielded using MCH incubation electrodes and placed at -5 to -3°C for 0.5–1.5 h.

[0042] Step S4: After the MCH incubation electrode has been shielded from non-specific binding sites, rinse it with buffer solution and store it in buffer solution to obtain an electrochemical biomolecular sensor based on a DNA molecular pendulum.

[0043] Example 4

[0044] This embodiment provides a method for constructing a biomarker protein electrochemical biomolecular sensor based on a DNA molecular pendulum, including the following steps:

[0045] (1) First, place the gold electrode (S = 0.2 mm) 2 The chamois was ultrasonically treated in ultrapure water for 3 minutes, and then polished with 0.3μm and 0.05μm alumina polishing powder for 5 minutes each. After the polishing was completed, it was rinsed with ultrapure water.

[0046] (2) The polished electrode needs further electrochemical cleaning and activation to remove surface contaminants. Cyclic voltammetry (CV) was used in 0.5 M H2SO4 at 0.05 V / s for 10 consecutive CV scans, with a potential range of -0.2 to 1.6 V (vsAg / AgCl).

[0047] (3) Incubate 5.0 μL of a 10 μM DNA double-stranded solution containing CEA (carcinoembryonic antigen) aptamers on the cleaned and activated gold electrode, then incubate at -20°C for 1 h. Remove the electrode, blow it clean, and use [Ru(NH3)6] to clean it. 3+The signal reduction molecule was used to incubate the electrode to modify the DNA molecule, and then placed at -4°C for 1 hour. After removing the gold electrode and cleaning it, the non-specific binding sites of the gold electrode were shielded by MCH and incubated at -4°C for 1 hour. Finally, the gold electrode was rinsed with Tris-HCl buffer and stored in 10 mM Tris-HCl buffer before use to prepare an electrochemical CEA sensor.

[0048] Example 5

[0049] This embodiment provides a method for constructing an RNA electrochemical biomolecular sensor based on the pendulum motion in a DNA molecule, including the following steps:

[0050] (1) First, place the gold electrode (S = 0.2 mm) 2 The microfiber was ultrasonically treated in ultrapure water for 3 minutes, and then polished on a microfiber cloth with 0.3μm and 0.05μm alumina polishing powder. After each polishing step (5 minutes), it was rinsed with ultrapure water.

[0051] (2) The polished electrode requires further electrochemical cleaning and activation to remove thiols / organic contaminants: Cyclic voltammetry (CV) was performed in 0.5 M H2SO4 at 0.05 V / s for 10 consecutive CV scans, with a potential range of -0.2 to 1.6 V (vsAg / AgCl).

[0052] (3) 5.0 μL of 10 μM DNA double-stranded solution containing RNA aptamers was incubated on the cleaned and activated gold electrode at -20°C for 4 hours. The electrode was removed and cleaned, and the gold electrode was incubated with ruthenium signal reducing molecules to modify the DNA molecules, and incubated at -4°C for 55 minutes. The gold electrode was then removed and cleaned again, and incubated with MCH at -20°C for 4 hours. Finally, the gold electrode was rinsed with Tris-HCl buffer and stored in 10 mM Tris-HCl buffer before use to prepare an RNA molecular pendulum electrochemical biosensor.

[0053] Example 6

[0054] This embodiment provides a method for constructing an electrochemical biological small molecule sensor based on a DNA molecular pendulum, including the following steps:

[0055] 1) First, place the gold electrode (S = 0.2 mm) 2 The chamois was ultrasonically treated in ultrapure water for 3 minutes, and then polished with 0.3μm and 0.05μm alumina polishing powder for 5 minutes each. After the polishing was completed, it was rinsed with ultrapure water.

[0056] (2) The polished electrode needs further electrochemical cleaning and activation to remove surface contaminants. Cyclic voltammetry (CV) was used in 0.5 M H2SO4 at 0.05 V / s for 10 consecutive CV scans, with a potential range of -0.2 to 1.6 V (vsAg / AgCl).

[0057] (3) Incubate 5.0 μL of 10 μM DNA double-stranded solution containing ATP (adenosine triphosphate) aptamers on the cleaned and activated gold electrode, then incubate at -20°C for 1 h. Remove the electrode, blow it clean, and use [Ru(NH3)6] 3+ The signal reduction molecule was used to incubate the electrode to modify the DNA molecule, and then placed at -4°C for 1 hour. After removing the gold electrode and cleaning it, the non-specific binding sites of the gold electrode were shielded by MCH and incubated at -4°C for 1 hour. Finally, the gold electrode was rinsed with Tris-HCl buffer and stored in 10 mM Tris-HCl buffer before use to prepare an electrochemical ATP sensor.

[0058] Example 7

[0059] This embodiment provides a method for detecting a disease marker protein (CEA), including the following steps:

[0060] (1) The test was performed using an electrochemical workstation with a three-electrode system. The Ag / AgCl electrode was used as the reference electrode, the platinum wire electrode was used as the auxiliary electrode, and the prepared biomolecular sensor was used as the working electrode. The working electrode was incubated with 5 μL of target molecules at concentrations of 0.01 ng / mL, 0.1 ng / mL, 1 ng / mL, 10 ng / mL, 100 ng / mL, and 1 μg / mL for 2 h and then placed in 10 mL of 10 mM Tris-HCl buffer at pH 7.4 for detection.

[0061] (2) The time-current method was used to detect it, with the potential range set to -0.1 to -0.5V and the running time to 0.2s.

[0062] (3) After the electrodes are placed, the current is measured every 2 seconds, and the working curve is plotted.

[0063] Example 8

[0064] This embodiment provides a method for detecting biomolecules (RNA), including the following steps:

[0065] (1) The test was performed using an electrochemical workstation with a three-electrode system. The saturated silver chloride electrode was used as the reference electrode, the platinum wire electrode was used as the auxiliary electrode, and the prepared biomolecular sensor was used as the working electrode. After incubating 5 μL of nucleotides (RNA) with concentrations of 1 nM, 10 nM, 100 nM, 1 μM, 10 μM and 100 μM for 2 h, the working electrode was placed in 10 mL of 10 mM Tris-HCl buffer at pH 7.4 for detection.

[0066] (2) The time-current method was used to detect it, with the potential range set to -0.1 to -0.5V and the running time to 0.2s.

[0067] (3) After the electrodes are placed, the current is measured every 2 seconds, and the working curve is plotted.

[0068] Example 9

[0069] This embodiment provides a method for detecting a biomolecule (ATP), including the following steps:

[0070] (1) The test was conducted using an electrochemical workstation with a three-electrode system. The saturated silver chloride electrode was used as the reference electrode, the platinum wire electrode was used as the auxiliary electrode, and the prepared biomolecular sensor was used as the working electrode. The working electrode was incubated with 5 μL of adenosine triphosphate (ATP) at concentrations of 1 nM, 10 nM, 100 nM, 1 μM, 10 μM and 100 μM for 2 h and then placed in 10 mL of 10 mM Tris-HCl buffer solution at pH 7.4 for detection.

[0071] (2) The time-current method was used to detect it, with the potential range set to -0.1 to -0.5V and the running time to 0.2s.

[0072] (3) After the electrodes are placed, the current is measured every 2 seconds, and the working curve is plotted.

[0073] Based on a DNA molecular pendulum-based electrochemical biomolecular sensor, we selected proteins (carcinoembryonic antigen, CEA), nucleotides (mRNA21), and small molecules (adenosine triphosphate, ATP) as target substances for detection. Figure 2 As shown, the timing current signal gradually decreases with increasing concentration, consistent with our theory, proving the successful construction of the sensing platform. This invention realizes the construction of the electrochemical biomolecular sensor and enables effective detection and precise quantitative analysis of biomolecules, including disease biomarker proteins, small molecules, and viruses.

[0074] This invention is based on the specific recognition and matching of biomolecules (disease biomarker proteins, RNA, etc.) and aptamers. Using a double-stranded DNA with an aptamer as the substrate, an Au electrode is fixed after modifying the DNA molecule with a ruthenium complex. In chronocurrent mode, an electric field opposite to the DNA molecule's pendulum is applied, causing it to oscillate and generate an electric signal, thereby achieving the detection of the target molecule. This invention realizes the construction of the electrochemical biomolecular sensor and enables the effective detection and precise quantitative analysis of biomolecules, including disease biomarker proteins, small molecules, and viruses. The constructed electrochemical biomolecular sensor has a wide detection range, is simple to construct, and exhibits high sensitivity and a low detection limit, making it significant for the detection of proteins, viruses, nucleic acids, and some small molecules.

[0075] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. An electrochemical biomolecular sensor based on a DNA molecular pendulum, characterized in that, The device includes a buffer solution, an electrode, and a DNA double-stranded pendulum, wherein the electrode and the DNA double-stranded pendulum are disposed in the buffer solution; a step voltage reduction signal molecule is inserted into the phosphate backbone of the DNA double-stranded pendulum; one end of the DNA double-stranded pendulum is connected to the electrode, and a target aptamer is connected to one of the single strands at the other end of the DNA double-stranded pendulum; the target aptamer is an aptamer that can specifically bind to CEA, PSA, RNA, or ATP; when the target aptamer on the DNA molecule pendulum binds to the target marker, its hydrodynamic diameter increases, and the time required for oscillation increases.

2. The electrochemical biomolecular sensor based on a DNA molecular pendulum according to claim 1, characterized in that: The step voltage reduction signal molecule is [Ru(NH3)6]. 3+ Signal reduction molecules.

3. The electrochemical biomolecular sensor based on a DNA molecular pendulum according to claim 1, characterized in that: The electrodes are gold electrodes, silver electrodes, silver chloride electrodes, and platinum electrodes.

4. The electrochemical biomolecular sensor based on a DNA molecular pendulum according to claim 2, characterized in that: The buffer solution is one or a mixture of two or more of the following: Tris-HCl solution or phosphate-buffered saline (PBS).

5. A detection method for non-therapeutic diagnostic purposes using the electrochemical biomolecular sensor based on a DNA molecular pendulum as described in claim 1, characterized in that: In chronocurrent mode, under the action of an applied step negative electric field, the DNA double-stranded pendulum swings toward the electrode. The step voltage reduces the signal molecule, generating a detectable Faraday current. When the target aptamer on the DNA double-stranded pendulum binds to the target marker, its hydrodynamic diameter increases, the time required for swinging increases, and the current decreases. By detecting the change in current, it can be determined whether the target marker has bound to the target aptamer on the DNA double-stranded pendulum, thereby realizing the detection of the target marker.

6. A method for constructing an electrochemical biomolecular sensor based on a DNA molecular pendulum as described in claim 1, comprising the following steps: Step S1: The electrode is ultrasonically treated in ultrapure water, then polished with aluminum oxide nanoparticles, and finally rinsed with ultrapure water. Step S2: Chemically clean and activate the polished electrode; Step S3: Incubate the DNA double-stranded lever containing the target aptamer on the cleaned and activated electrode, remove the electrode and blow it clean, then use [Ru(NH3)6] to clean it. 3+ The signal reduction molecule is incubated with the electrode to modify the DNA molecule; then the electrode is removed, blown clean, and incubated with MCH to shield non-specific binding sites. Step S4: After the MCH incubation electrode has been shielded from non-specific binding sites, rinse it with buffer solution and store it in buffer solution to obtain an electrochemical biomolecular sensor based on a DNA molecular pendulum.

7. The method for constructing an electrochemical biomolecular sensor based on a DNA molecular pendulum according to claim 6, characterized in that: In step S2, the polished and cleaned electrode is activated in H2SO4 using cyclic voltammetry. CV scans are performed continuously at 0.03-0.7 V / s, with a potential range of -0.2 to 1.6 V.

8. The method for constructing an electrochemical biomolecular sensor based on a DNA molecular pendulum according to claim 7, characterized in that: In step S3, 4.0-6.0 μL of 8-12 μM DNA double-stranded pendulum containing the target aptamer is incubated on the cleaned and activated electrode, and then placed in a -20°C freezer for 0.5-1.5 h.

9. The method for constructing an electrochemical biomolecular sensor based on a DNA molecular pendulum according to claim 8, characterized in that: In step S3, [Ru(NH3)6] is used. 3+ Signal reduction molecule incubation electrode to modify DNA molecules, placed in an environment of -5 to -3℃ for 0.5 to 1.5 h; MCH incubation electrode to shield non-specific binding sites, placed in an environment of -5 to 4℃ for 0.5 to 1.5 h.

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