阅读说明：本技术 自加热生物传感器芯片及其制备方法、病毒检测方法 (Self-heating biosensor chip, preparation method thereof and virus detection method ) 是由 刘俊江 王焕明 王艳丽 许智 于 2021-09-16 设计创作，主要内容包括：一种自加热生物传感器芯片及其制备方法、病毒检测方法,属于生物检测领域。自加热生物传感器芯片包括衬底、二氧化硅层、检测单元以及加热电极。二氧化硅层形成并覆盖于衬底的支撑面,检测单元包括：分别形成于二氧化硅层的承载面的碳膜以及两个金属电极,两个金属电极分别与碳膜的两端电连接,碳膜被配置为接触样品溶液,碳膜具有用于修饰生物大分子的修饰面。加热电极形成于衬底内,加热电极的加热面与二氧化硅层抵接以通过热传导加热碳膜。加热电极的工作电压被配置为20～35mV,位于碳膜与加热电极之间的二氧化硅层的厚度为150～400nm。其能够解决生物传感器无法重复利用的问题,以提高利用率以及降低检测成本。(A self-heating biosensor chip, a preparation method thereof and a virus detection method belong to the field of biological detection. The self-heating biosensor chip includes a substrate, a silicon dioxide layer, a detection unit, and a heating electrode. The silicon dioxide layer is formed and covered on the supporting surface of the substrate, and the detection unit comprises: the two metal electrodes are respectively and electrically connected with two ends of the carbon film, the carbon film is configured to contact the sample solution, and the carbon film is provided with a modification surface used for modifying the biomacromolecule. The heating electrode is formed in the substrate, and a heating surface of the heating electrode is in contact with the silicon dioxide layer to heat the carbon film by thermal conduction. The working voltage of the heating electrode is set to 20-35 mV, and the thickness of the silicon dioxide layer between the carbon film and the heating electrode is 150-400 nm. The biosensor can solve the problem that the biosensor cannot be repeatedly used, so that the utilization rate is improved, and the detection cost is reduced.)

1. A self-heating biosensor chip, comprising:

a substrate having a support surface;

the silicon dioxide layer is formed and covers the supporting surface, and the silicon dioxide layer is provided with a bearing surface far away from the substrate;

at least one detection unit, each said detection unit comprising: the device comprises a carbon film and two metal electrodes, wherein the carbon film and the two metal electrodes are respectively formed on the bearing surface, the two metal electrodes are respectively electrically connected with two ends of the carbon film, the carbon film is configured to contact a sample solution, and the carbon film is provided with a modification surface used for modifying biological macromolecules capable of detecting action objects in the sample solution; and

a heating electrode formed in the substrate, a heating surface of the heating electrode abutting against the silicon dioxide layer to heat the carbon film by thermal conduction;

wherein the working voltage of the heating electrode is set to be 20-35 mV, and the thickness of the silicon dioxide layer between the carbon film and the heating electrode is 150-400 nm.

2. The self-heating biosensor chip of claim 1, wherein the carbon material in the carbon film comprises at least one of graphene, fullerene, and carbon nanotubes;

optionally, the carbon material in the carbon film is graphene.

3. The self-heating biosensor chip of claim 1, wherein said modified surface is modified with said biomacromolecule, said biomacromolecule being a viral antigen, a viral antibody or a nucleic acid probe.

4. The self-heating biosensor chip according to claim 1, wherein the number of the detection units is at least two, and any two adjacent detection units are arranged at intervals.

5. The self-heating biosensor chip according to claim 4, wherein the number of the heating electrodes corresponds to the number of the detection units, each heating electrode is disposed at the orthographic projection of the detection unit on the supporting surface, and any two adjacent heating electrodes are disposed at intervals.

6. The self-heating biosensor chip according to any one of claims 1 to 5, further comprising a gate electrode disposed on the carrying surface and configured to adjust a field effect of each of the carbon films.

7. A method for preparing a self-heating biosensor chip, comprising:

providing a substrate;

depositing a heating electrode on the supporting surface of the substrate, then depositing a silicon dioxide layer on the rest supporting surface and the surface of the heating electrode, and etching the silicon dioxide layer to expose a bonding pad of the heating electrode;

forming a patterned carbon film on the bearing surface by taking one surface of the silicon dioxide layer, which is far away from the substrate, as the bearing surface, depositing two metal electrodes which are respectively electrically connected with two ends of the carbon film on the bearing surface, and packaging the bearing surface and exposing the carbon film, a bonding pad of each metal electrode and a bonding pad of the heating electrode;

and modifying the modification surface of the exposed carbon film with a biological macromolecule, wherein the biological macromolecule is a virus antigen, a virus antibody or a nucleic acid probe.

8. The method of claim 7, wherein after depositing the two metal electrodes on the carrying surface, the method further comprises depositing a gate electrode on the carrying surface and exposing the gate electrode after the encapsulating, the gate electrode being configured to modulate a field effect of the carbon film.

9. The method according to claim 7, wherein the biomacromolecule is an antibody, and the step of modifying the biomacromolecule with the carbon membrane comprises:

functionalizing the surface of the carbon film by adopting a PBASE/ethanol solution to form a PBASE/carbon film;

spotting an antibody/PBS solution on the PBASE/carbon film, drying, blocking PBASE which is not combined with the antibody by using a blocking solution, and washing.

10. A method for detecting a virus, comprising:

obtaining a self-heating biosensor chip according to any one of claims 1 to 6 or a self-heating biosensor chip prepared by the preparation method according to any one of claims 7 to 9;

contacting a sample solution with the carbon film, detecting currents at two ends of the carbon film by using the two metal electrodes, and determining that the sample solution contains viruses which specifically react with biological macromolecules if the currents at two ends of the carbon film change;

heating the carbon film using the heating electrode to desorb the virus bound to the bio-macromolecule.

Technical Field

The application relates to the field of biological detection, in particular to a self-heating biosensor chip, a preparation method thereof and a virus detection method.

Background

The bioelectricity sensor is different from the traditional biological detection technology, converts the detection of viruses into the detection of electric signals, and is quicker and more efficient. However, in most cases, biosensors such as graphene biosensors are applied only once, and have low recycling rate, resulting in high production and detection costs.

Disclosure of Invention

The application provides a self-heating biosensor chip, a preparation method thereof and a virus detection method, which can solve the problem that a biosensor cannot be reused.

The embodiment of the application is realized as follows:

in a first aspect, the present examples provide a self-heating biosensor chip comprising a substrate, a silicon dioxide layer, at least one detection cell, and a heating electrode.

The substrate has a support surface.

The silicon dioxide layer is formed and covered on the supporting surface, and the silicon dioxide layer is provided with a bearing surface far away from the substrate.

Each detection unit includes: the biological macromolecule heating device comprises a carbon film and two metal electrodes, wherein the carbon film and the two metal electrodes are respectively formed on a bearing surface, the two metal electrodes are respectively electrically connected with two ends of the carbon film, the carbon film is configured to be in contact with a sample solution, the carbon film is provided with a modification surface, the modification surface is used for modifying a biological macromolecule heating electrode capable of detecting an action object in the sample solution and is formed in a substrate, and the heating surface of the heating electrode is abutted to a silicon dioxide layer so as to heat the carbon film through heat conduction.

Wherein the working voltage of the heating electrode is set to be 20-35 mV, and the thickness of the silicon dioxide layer between the carbon film and the heating electrode is 150-400 nm.

In the implementation process, after the self-heating biosensor chip is detected, based on the mutual matching of the heating electrode with the working voltage of 20-35 mV and the silicon dioxide layer with the thickness of 150-400 nm between the carbon film and the heating electrode, the carbon film is ensured to have proper temperature, viruses and residual biological solution combined with biological macromolecules on the surface of the carbon film can be removed, and the biological macromolecules after the viruses are removed are effectively prevented from being inactivated, so that the self-heating biosensor chip is recycled, and the detection cost is effectively reduced; and because the carbon film is the ultrahigh thermal conductivity material, the voltage that needs is very big if the carbon film is heated directly, has not only increased the consumption and damaged sensor itself easily, and this application utilizes the setting of the silica layer that the thickness is 150 ~ 400nm between carbon film and the heating electrode, guarantees that the operating voltage of heating the carbon film to preset temperature is only 20 ~ 35mV, effectively reduces the energy consumption.

In one possible embodiment in combination with the first aspect, the carbon material in the carbon film comprises at least one of graphene, fullerene, and carbon nanotube.

Optionally, the carbon material in the carbon film is graphene.

The sensitivity of the chip prepared from the graphene to viruses is good.

In a possible embodiment in combination with the first aspect, the modification surface is modified with a biological macromolecule, the biological macromolecule being a viral antigen, a viral antibody or a nucleic acid probe.

With reference to the first aspect, in one possible embodiment, the number of the detection units is at least two, and any two adjacent detection units are arranged at intervals.

In the implementation process, the number of the detection units is at least two, so that the same biological macromolecules can be modified on the carbon film in each detection unit according to actual requirements to carry out parallel experiments or contrast experiments, or different biological macromolecules can be modified on the carbon film in each detection unit to detect different sample solutions simultaneously, and the detection efficiency is improved.

With reference to the first aspect, in a possible implementation, the number of the heating electrodes corresponds to that of the detection units, each heating electrode is arranged at an orthographic projection of the detection unit on the bearing surface, and any two adjacent heating electrodes are arranged at intervals.

In the implementation process, whether the heating electrode corresponding to each detection unit works or not can be controlled independently, and the influence on another adjacent detection unit when the heating electrode corresponding to one detection unit works can be effectively avoided by combining the specific arrangement position of each heating electrode.

With reference to the first aspect, in one possible embodiment, the self-heating biosensor chip further comprises a gate electrode disposed on the carrying surface and configured to modulate the field effect of each carbon film.

The field effect of each carbon film is adjusted by the gate electrode, thereby adjusting the current through the carbon film.

With reference to the first aspect, in a possible implementation, the substrate is a silicon wafer, and the material of the silicon dioxide layer is silicon dioxide.

In a second aspect, the present application provides a method of preparing a self-heating biosensor chip, comprising:

a heating electrode is deposited on the supporting surface of the substrate, then a silicon dioxide layer is deposited on the rest supporting surface and the surface of the heating electrode, and the silicon dioxide layer is etched to expose the bonding pad of the heating electrode.

And taking one surface of the silicon dioxide layer, which is far away from the substrate, as a bearing surface, forming a patterned carbon film on the bearing surface, depositing two metal electrodes which are respectively and electrically connected with two ends of the carbon film on the bearing surface, packaging the bearing surface, and exposing the carbon film, the bonding pad of each metal electrode and the bonding pad of the heating electrode.

And modifying the exposed carbon film with biological macromolecules, wherein the biological macromolecules are virus antigens, virus antibodies or nucleic acid probes.

Wherein the working voltage of the heating electrode is set to be 20-35 mV, and the thickness of the substrate between the carbon film and the heating electrode is 150-400 nm.

The preparation method is simple to operate, and the prepared self-heating biosensor chip can be repeatedly utilized, so that the detection efficiency is effectively improved, and the cost is reduced.

In combination with the second aspect, in a possible embodiment, after depositing the two metal electrodes on the carrying surface, the preparation method further comprises depositing a gate electrode on the carrying surface, and exposing the gate electrode after encapsulation, the gate electrode being configured to modulate the field effect of the carbon film.

In combination with the second aspect, in one possible embodiment, the biomacromolecule is an antibody, and the step of carbon membrane modification of the biomacromolecule comprises:

functionalizing the surface of the carbon film by using a PBASE/ethanol solution to form the PBASE/carbon film.

Spotting an antibody/PBS solution on the PBASE/carbon film, drying, blocking PBASE which is not combined with the antibody by using a blocking solution, and washing.

By adopting the modification method, the antibody is stably combined on the carbon film through the PBASE, the PBASE which is not combined with the antibody is sealed at the same time, the biological macromolecule can be ensured to be specifically combined with the antigen, and the PBASE is taken as a connecting molecule, so that the antibody can be stably combined on the carbon film within the heating temperature range and cannot be separated from the carbon film.

In a second aspect, the present application provides a virus detection method, including:

the self-heating biosensor chip provided by the first aspect of the present application or the self-heating biosensor chip prepared by the preparation method provided by the second aspect is obtained.

And (3) contacting the sample solution with the carbon film, detecting the current at two ends of the carbon film by using two metal electrodes, and determining that the sample solution contains the virus which is specifically reacted with the biological macromolecules if the current at two ends of the carbon film changes.

The carbon film is heated by the heating electrode to desorb the virus bound to the bio-macromolecules.

The detection mode is simple, and the virus combined with the biomacromolecule can be desorbed by heating the heating electrode after the detection result comes out, and the biomacromolecule still keeps activity, so that the repeated use is realized.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings that are required to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present application and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained from the drawings without inventive effort.

FIG. 1 is a schematic structural view of a self-heating biosensor chip;

FIG. 2 is a schematic structural diagram of a chip body;

FIG. 3 is a schematic view of the structure in the direction A-A in FIG. 2;

FIG. 4 is a schematic structural diagram of a detecting unit disposed on the surface of a silicon dioxide layer;

fig. 5 is a schematic diagram of the current through graphene as a function of time provided in experimental example 1;

FIG. 6 is a schematic diagram of the current through graphene under dry-fire conditions as a function of time as provided in test example 1;

FIG. 7 is a schematic diagram of the detection sensitivity of a self-heating biosensor chip;

fig. 8 is a schematic diagram of the current through graphene as a function of time provided in experimental example 3.

Icon: 10-self-heating biosensor chip; 100-a chip body; 110-a substrate; 120-a silicon dioxide layer; 131-carbon film; 133-a metal electrode; 140-a heating electrode; 150-a gate electrode; 170-an encapsulation layer; a 200-PDMS layer; 210-sample addition tank.

Detailed Description

Embodiments of the present application will be described in detail below with reference to examples, but those skilled in the art will appreciate that the following examples are only illustrative of the present application and should not be construed as limiting the scope of the present application. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.

Example 1

Referring to fig. 1 and 2, a self-heating biosensor chip 10 includes a chip body 100 and a PDMS layer 200.

Referring to fig. 2, 3 and 4, the chip body 100 includes a substrate 110, a silicon dioxide layer 120, a detection unit, a heating electrode 140, a gate electrode 150 and a packaging layer 170.

Wherein, the substrate 110 has a supporting surface, the silicon dioxide layer 120 is formed and covered on the supporting surface, and the silicon dioxide layer 120 has a carrying surface far away from the substrate 110; the gate electrode 150 and the detecting unit are respectively disposed on the carrying surface, and the gate electrode 150 is configured to adjust a field effect of each carbon film 131. The detection unit includes a carbon film 131 and two metal electrodes 133 respectively formed on the carrying surface, the two metal electrodes 133 are respectively electrically connected with two ends of the carbon film 131, the carbon film 131 is configured to contact and detect an action object in the sample solution, the carbon film 131 has a modification surface for modifying a bio-macromolecule capable of detecting the action object in the sample solution, the heating electrode 140 is formed in the substrate 110, and a heating surface of the heating electrode 140 abuts against the silicon dioxide layer 120 to heat the carbon film 131 through heat conduction.

The above arrangement can utilize the contact between the sample solution and the carbon film 131, and the two metal electrodes 133 to detect whether the current at the two ends of the carbon film 131 changes, so as to determine whether the sample solution contains the virus, which is the target of specific reaction with the biological macromolecule, and after the determination is completed, the heating electrode 140 can be used to heat the carbon film 131 by in-situ heat conduction, so as to remove the virus and the biological solution on the surface of the carbon film 131 combined with the biological macromolecule under the premise of low power consumption.

The silicon dioxide layer 120 has insulation properties and thermal conductivity. In order to avoid the failure of the biomacromolecules on the carbon film 131 caused by the overlarge temperature in the actual use process, the technical problem of the application cannot be solved due to the undersize, and the problem can be effectively solved by controlling the thickness of the silicon dioxide layer 120 between the carbon film 131 and the heating electrode 140 and the mutual matching of the working voltage of the heating electrode 140.

The operating voltage of the heating electrode 140 is set to 20 to 35mV, the thickness of the silicon dioxide layer 120 between the carbon film 131 and the heating electrode 140 is 150 to 400nm, and the thickness of the silicon dioxide layer 120 between the carbon film 131 and the heating electrode 140 is 150nm, 160nm, 170 nm, 180nm, 190nm, 200nm, 250nm, 280nm, 300nm, 350nm, or 400nm, for example. Based on specific voltage, the position relation of the heating electrode 140 and the carbon film 131, and the matching of the silicon dioxide layer 120 and the thickness thereof, the inactivation of the virus-removed biological macromolecules can be effectively avoided after viruses and residual biological solution combined with the biological macromolecules on the surface of the carbon film 131 are removed, so that the reutilization of the self-heating biosensor chip is realized, and the energy consumption is effectively reduced.

In the present embodiment, the operating voltage of the heating electrode 140 was set to 35mV, and the thickness of the silicon dioxide layer 120 between the carbon film 131 and the heating electrode 140 was 200 nm.

The biological macromolecules can be prepared or purchased according to viruses which need to be detected actually, and meanwhile, the viruses include but are not limited to new coronavirus, influenza virus and the like, and the biological macromolecules can be limited by those skilled in the art according to actual needs and are not limited herein.

For example, in this example, the biomacromolecule is the novel coronavirus antibody Mab Mouse anti-Covid-19N.

The substrate 110 is used for supporting a silicon dioxide layer 120 formed on a supporting surface, and in the embodiment, the substrate 110 is a silicon wafer, and the thickness and size thereof can be selected according to actual requirements.

In order to facilitate the control of the thickness of the silicon dioxide layer 120 between the carbon film 131 and the heating electrode 140, as shown in fig. 3, the supporting surface is provided with a groove, the heating electrode 140 is deposited in the groove, and the heating surface faces the opening of the groove and is flush with the supporting surface, that is, the depth of the groove is the same as the thickness of the heating electrode 140, for example, in this embodiment, the depth of the groove is 50nm, and the thickness of the heating electrode 140 is 50 nm. The silicon dioxide layer 120 is formed to cover the supporting surface (the supporting surface without the groove) and the heating surface, so that the heating surface can transfer heat to the silicon dioxide layer 120 by means of heat conduction.

The number of the heating electrodes 140 is at least one, such as one, two, or three, etc., and the number of the sensing units is at least one, such as one, two, or three, etc.

In the first arrangement provided by the present application, the detecting units correspond to the number of the heating electrodes 140 one by one, and each heating electrode 140 is disposed at the orthographic projection of the detecting unit on the supporting surface. That is, the heating electrodes 140 corresponding to each detection unit are used to individually heat the corresponding carbon film 131, so that the use is more flexible, and in the actual use process, when only some of the heating electrodes 140 corresponding to the detection units are used to work to remove viruses bound on the corresponding carbon film 131 and/or dry the carbon film 131, the influence on the idle bio-macromolecules on the carbon film 131 adjacent to the heating electrodes can be avoided.

It should be noted that, in order to ensure good heating effect, the area of the heating electrode 140 is equal to or slightly larger than the area of the orthographic projection.

In some optional examples in the case of the first setting, the number of the detection units and the number of the heating electrodes 140 are each one.

In other optional examples in the case of the first arrangement, the number of the detection units and the number of the heating electrodes 140 are at least two, for example, the number of the detection units is two, three, five, and the like. Meanwhile, the same biological macromolecules can be modified on the carbon films 131 in different detection units according to actual requirements so as to perform parallel experiments or contrast experiments, or different biological macromolecules can be modified on the carbon films 131 in different detection units so as to detect different sample solutions simultaneously, so that the detection efficiency is improved.

Optionally, any two adjacent detection units are arranged at intervals, and any two adjacent heating electrodes 140 are arranged at intervals, so as to avoid that the heating electrode 140 corresponding to one detection unit affects another detection unit adjacent to the detection unit when operating. It should be noted that the distance between any two adjacent detection units and the distance between any two adjacent heating electrodes 140 can be set according to the actual chip layout.

For example, in the embodiment, the number of the detection units and the number of the heating electrodes 140 are three, and the three heating electrodes 140 and the three detection units are respectively arranged in an array.

In the second arrangement, the number of detection cells is greater than the number of heater electrodes 140.

That is, the number of the sensing units is at least two, and the number of the heating electrodes 140 is at least one. At this time, the adjacent two carbon films 131 are arranged at intervals, and the heating electrode 140 is at an orthographic projection covering each detection unit on the support surface.

At this time, the same biological macromolecules can be modified on the carbon films 131 in different detection units according to actual requirements to perform parallel experiments or control experiments, or different biological macromolecules can be modified on the carbon films 131 in different detection units to detect different sample solutions simultaneously, so that the detection efficiency is improved. However, it should be noted that, in this case, for the purpose of use, it is recommended that a plurality of detection units be heated together after use for recycling.

The carbon material in the carbon film 131 in the detection unit includes at least one of graphene, fullerene and carbon nanotube, for example, the carbon material is graphene, fullerene or carbon nanotube, or the like, or a mixture of any two of the graphene, fullerene and carbon nanotube, which are all ultra-high thermal conductivity materials, and have repairability and excellent electrical conductivity, and can be used as a test electrode in this application.

In this embodiment, the carbon material is graphene, and the sensitivity of the self-heating biosensor chip 10 is better.

In order to facilitate subsequent connection with an external power source and inspection operation, the silicon dioxide layer 120 is provided with a first opening (not shown) for exposing the pad of the heating electrode 140.

The encapsulation layer 170 is formed on the carrying surface to encapsulate the silicon dioxide layer 120, the carbon film 131, the first metal electrode 133, the second metal electrode 133 and the gate electrode 150.

Wherein the encapsulation layer 170 is provided with third openings for exposing the gate electrode 150 and the pad thereof, the pad of the first metal electrode 133, the pad of the second metal electrode 133, and the pad of the heating electrode 140, respectively, and a second opening for exposing the carbon film 131, wherein the third opening for exposing the pad of the heating electrode 140 communicates with the first opening.

The PDMS layer 200 is disposed on a surface of the encapsulation layer 170 away from the silicon dioxide layer 120. PDMS (polydimethylsiloxane) has the advantages of excellent flexibility, transparency, fluidity and biocompatibility, and its thickness can be set according to practical requirements, for example, the thickness of the PDMS layer 200 is 2mm in this embodiment.

PDMS layer 200 is equipped with the three fourth opening of second opening one-to-one, three fourth opening interval arrangement, every fourth open-ended cross sectional area is greater than the second open-ended cross sectional area and the projection of fourth opening at the loading face covers the second opening completely, the fourth opening communicates with the second opening and forms the ladder groove as sample adds groove 210, sample adds groove 210 utilizes the cross sectional area of fourth opening to be greater than the cross sectional area of second opening so that add the sample solution of detection, simultaneously because carbon film 131 highly is less than encapsulating layer 170, therefore it has sufficient degree of depth in order to avoid in-service use sample solution spill scheduling problem.

The present application provides a method for preparing the self-heating biosensor chip 10, which comprises the following steps:

s1, providing the substrate 110.

S2, depositing the heater electrode 140 on the supporting surface of the substrate 110, then depositing the silicon dioxide layer 120 on the remaining supporting surface and the surface of the heater electrode 140, and etching the silicon dioxide layer 120 to expose the pad of the heater electrode 140.

S3, forming a patterned carbon film 131 on the carrying surface with the surface of the silicon dioxide layer 120 away from the substrate 110 as the carrying surface, depositing two metal electrodes 133 on the carrying surface, which are electrically connected to two ends of the carbon film 131, respectively, and then depositing a gate electrode 150 on the carrying surface, which is configured to adjust the field effect of the carbon film 131, to obtain the chip body 100, then encapsulating the carrying surface with an encapsulation layer 170 and exposing the carbon film 131, a pad of each metal electrode 133, a pad of the heating electrode 140, and a pad of the gate electrode 150, and then encapsulating the encapsulation layer 170 with a PDMS layer 200 and exposing the carbon film 131.

After the carrying surface is encapsulated, the exposed carbon film 131 is modified with a bio-macromolecule, which in this embodiment is a virus antibody.

The carbon membrane 131 modification of the biomacromolecule comprises the following steps:

functionalizing the surface of the carbon film 131 by using a PBASE/ethanol solution to form a PBASE/carbon film 131; and (3) spotting the antibody/PBS solution on the PBASE/carbon film 131, drying, then blocking the PBASE which is not combined with the antibody by using a blocking solution, and washing.

In the embodiment, a 2mM PBASE/ethanol solution is dripped on the surface of graphene for 2 hours, then the graphene is washed by ethanol, and dried by an air gun, so that PBASE/graphene is obtained. Dropwise adding an antibody/PBS solution on the PBASE/graphene for 4h, washing with the PBS solution, blow-drying with an air gun, dropwise adding 1M ethanolamine/PBS for 1h to seal the PBASE, and then washing to obtain the sealed antibody/PBASE/graphene.

The manufacturing steps of the chip body 100 may be performed using a photolithography process.

In this embodiment, the chip body 100 is prepared by the following preparation method:

spin-coating photoresist on the supporting surface of the silicon wafer, carrying out ultraviolet exposure to obtain a patterned groove etching area, etching the patterned groove etching area by using an inductively coupled plasma device to obtain a groove with the etching depth of 50nm, depositing a heating electrode 140 with the thickness of 50nm in the groove, removing the photoresist and leaving the heating electrode 140.

A silicon dioxide layer 120 with a thickness of 200nm is deposited on the support surface, and the prepared heating electrode 140 is covered with the silicon dioxide layer 120. Then, a photoresist is coated on the carrying surface of the silicon dioxide layer 120 in a spinning manner, a pad exposure region corresponding to the pad of the heating electrode 140 is formed by ultraviolet exposure, then, the pad exposure region is etched to expose the pad of the heating electrode 140, and the photoresist is removed.

Transferring graphene to a bearing surface, then spin-coating photoresist on the surface of the graphene, carrying out ultraviolet exposure, and etching the exposed graphene by using a plasma cleaning machine to obtain patterned graphene.

And then, spin-coating photoresist on the surfaces of the graphene and the bearing surface, performing ultraviolet exposure to form two metal electrode deposition areas corresponding to two ends of the graphene film, and removing the photoresist after depositing metal electrodes 133 electrically connected with the graphene in each metal electrode deposition area.

Spin-coating photoresist on the bearing surface, performing ultraviolet exposure to form a gate electrode deposition region, depositing a gate electrode 150 on the gate electrode deposition region, removing the photoresist, and then encapsulating the bearing surface with an encapsulation layer 170 and exposing the carbon film 131, the pad of each metal electrode 133, and the pads of the heater electrode 140 and the gate electrode 150.

The present application also provides a virus detection method using the self-heating biosensor chip 10, the virus detection method including:

the sample solution is brought into contact with the carbon film 131, and the current at both ends of the carbon film 131 is detected by the two metal electrodes 133, and if the current at both ends of the carbon film 131 changes, it is determined that the sample solution contains a virus specifically reacting with the biological macromolecule.

After the test is completed, the carbon film 131 is heated by the heating electrode 140 through heat conduction to desorb the virus bound to the bio-macromolecule.

Example 1 was repeated to prepare 5 self-heating biosensor chips 10 as a first self-heating biosensor chip, a second self-heating biosensor chip, a third self-heating biosensor chip, a fourth self-heating biosensor chip and a fifth self-heating biosensor chip, respectively, in the same manner, to perform the following test examples.

Test example 1

The sample addition bath corresponding to any of the detection units of the first self-heating biosensor chip prepared in example 1 was added dropwise with the SARS-CoV-2 nucleomapped-His recombinant Protein antigen, and the operation of the heating electrode corresponding to the sample addition bath was started at 750s with the operating voltage of the heating electrode being 35mV, so that a graph showing the change with time of the current passing through graphene as shown in FIG. 5 was obtained.

According to fig. 5, it can be seen that the antibody on the surface of the graphene is combined with the antigen within 0-750s, the heating electrode is powered on at 750s, the antigen is gradually desorbed with the increase of time, the current starts to rise at the moment, and is finally desorbed completely, and the current curve is stable; after the antigen is added again, the antibody binds to the antigen rapidly, resulting in a drop in current and a return to the previous value, indicating that the antibody is not inactivated and the antigen remains bound to the antibody.

And (3) repeating the test by using the remaining two detection units to obtain a similar schematic diagram of the change of the current passing through the graphene along with the time, which shows that the working voltage of the heating electrode is 35mV, the antibody is not inactivated, and the antigen can still be combined with the antibody.

Meanwhile, in the experimental process, whether the dry-burning condition occurs or not can be judged through a change graph of the current passing through the graphene along with the time. As shown in FIG. 6, the SARS-CoV-2 nucleocapside-His recombinant Protein antigen was dropped into the sample addition well corresponding to any of the detection cells of the second self-heating biosensor chip prepared in example 1, and the operation of the heater electrode corresponding to the sample addition well was started at 1030s with a heater electrode operating voltage of 35 mV.

As can be seen from fig. 5 and fig. 6, in the antibody + antigen phase, the current is relatively stable, and then no additional antigen is added after the heating electrode is operated (the heating electrode is started at about 1030s in fig. 6), and it can be seen that the current is instantaneously reduced after the current is increased to a stable state due to the antigen being separated from the antibody, indicating that dry burning occurs at this time. That is, whether dry burning occurs or not can be judged by using the schematic diagram of the change of the current passing through the graphene along with the time.

Test example 2

The third self-heating biosensor chip prepared in example 1 was subjected to sensitivity detection, which comprises the following steps: 1 XPBS solution without antigen (SARS-CoV-2Nucleocapsid-His recombinant Protein) is added dropwise, and then the solution is added with 10 concentration^-18～10^-8The 1x PBS solution of the antigen (SARS-CoV-2Nucleocapsid-His recombinant Protein) was subjected to a change in current depending on the concentration of the antigen in the dropwise added solution, and the sensitivity was obtained by utilizing the trend of the current change, as shown in FIG. 7.

As can be seen from FIG. 7, the third self-heating biosensor chip had an antigen concentration of 10 even when the concentration was measured^-18M also has higher sensitivity. The above-described experiment was repeated for the fourth self-heating biosensor chip prepared in example 1, and similar results were obtained.

Test example 3

After 1nM of SARS-CoV-2 nucleomapped-His receptor Protein was dropped into the sample addition bath corresponding to any of the detection units of the fifth self-heating biosensor chip prepared in example 1, the heating electrode corresponding to the sample addition bath was operated with an energization voltage of 45mV, and a schematic diagram of the change with time of the current passing through the graphene as shown in fig. 8 was obtained.

From FIG. 8, it can be seen that the heating temperature is high when the working voltage of the heating electrode is 45mV, and after heating, the antibody is inactivated, and after adding the antigen, the current cannot return to the original value.

In summary, the self-heating biosensor chip, the preparation method thereof and the virus detection method can solve the problem that the biosensor cannot be reused, and meanwhile, the repeated use and low power consumption effectively reduce the detection cost.

The foregoing is merely exemplary of the present application and is not intended to limit the present application, which may be modified or varied by those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present application shall be included in the protection scope of the present application.