A microfluidic integrated biosensor system for collecting and detecting respiratory gas condensate

Abstract

The application provides a microfluidic integrated biosensor system integrating breath gas condensate collection and detection, comprising: a breath gas condensate collector based on thermoelectric cooling, which comprises: a hose, a condensate collection tube, a cooling trap aluminum block, a Peltier element and an MCU cooler; a microfluidic integrated on-chip gate graphene field effect transistor biosensor system, which comprises: a microfluidic chip and an on-chip gate graphene field effect transistor biosensor; a power supply and a signal acquisition and control module; wherein the exhaled gas condenses after entering the condensate collection tube, and then flows into the microfluidic chip and the surface of the on-chip gate graphene field effect transistor biosensor, and the signal acquisition and control module is used for collecting the biosensor signal, so that the detection of biomarkers in the breath gas condensate is realized. The application can realize the collection and detection of EBC at the same time, which not only reduces the work burden of the sampling personnel, but also avoids the risk of sample contamination.

Description

Technical Field

[0001] This invention relates to the field of biosensor technology, and more specifically to a microfluidic integrated biosensor system that integrates the collection and detection of respiratory gas condensates. Background Technology

[0002] Studies have shown that infectious SARS-CoV-2 exists in the exhaled breath condensate (EBC) of individuals infected with COVID-19, making EBC samples a promising candidate for detection and screening of the novel coronavirus. Currently, EBC sample collection devices mainly fall into several categories, with commercially available devices including the RTube from Respiratory Research in the United States. TM Exhaled blood gas (EBC) sample collection utilizes systems such as the TurboDECCS system from Medivac (Italy), as well as self-made systems using ice baths, frozen aluminum baths, and thermoelectric coolers. However, these systems only collect EBC samples; samples typically need to be transferred before analysis and testing. This increases the risks and workload for sampling personnel, as well as the risk of sample contamination. In recent years, companies like Exhalation Technology (UK) have developed integrated systems for collecting respiratory gas condensates and detecting physicochemical indicators (hydrogen peroxide and carbon dioxide, etc.), but to date, no integrated system for detecting biomolecules in respiratory gas condensates has been developed.

[0003] Under normal respiratory conditions, the viral load of SARS-CoV-2 in exhaled droplets entering the EBC ranged from 3.92 × 10⁻⁶. -1 copy / mL up to 5.10×10 4 The sensitivity level is around 1000 ppm, requiring biosensors with very high sensitivity to detect the novel coronavirus in respiratory gas condensates. Among the many diagnostic methods currently available, field-effect transistor (FET)-based biosensors not only possess this level of sensitivity but also offer several other advantages, including the ability to perform high-sensitivity and instantaneous measurements with small amounts of analyte. FET-based biosensors are considered to have promising applications in clinical diagnostics, point-of-care testing, and field testing. Graphene, due to its exceptional properties, including high electronic conductivity and high carrier mobility, is used in various sensing platforms. Graphene-based FET biosensors can detect changes in their surface and provide an optimal sensing environment for ultra-sensitive and low-noise detection, showing promising applications in sensitive immunodiagnostics. Summary of the Invention

[0004] The purpose of the present application is to provide a microfluidic integrated biosensor system integrating breath gas condensate collection and detection, so as to solve the problem of lack of integrated detection system for biomolecules in breath gas condensate in the prior art.

[0005] In order to solve the above technical problems, the present application adopts the following technical solutions:

[0006] The present application provides a microfluidic integrated biosensor system integrating breath gas condensate collection and detection, comprising: a breath gas condensate collector based on thermoelectric cooling, comprising: a hose, a condensate collection tube connected with the hose, a cooling trap aluminum block sleeved outside the condensate collection tube, a thermoelectric cooler Peltier element closely arranged on one side of the cooling trap aluminum block, and an MCU cooler; a microfluidic integrated on-chip gate graphene field effect transistor biosensor system, comprising: a microfluidic chip and an on-chip gate graphene field effect transistor biosensor bonded by plasma, the on-chip gate graphene field effect transistor biosensor having a graphene biosensor channel, on which streptavidin or protein G capable of binding with coronavirus antibodies is modified; a power supply and a signal acquisition and control module; wherein the exhaled gas enters the condensate collection tube through the hose and condenses to form condensate which flows into the microfluidic chip and the surface of the on-chip gate graphene field effect transistor biosensor, and the biosensor signal is collected by the signal acquisition and control module, so as to realize the detection of biomarkers in breath gas condensate.

[0007] The condensate collection tube comprises: a breath gas introduction pipe, a condensing pipe body connected with the breath gas introduction pipe, a capillary one-way upper stop valve arranged above the condensing pipe body, and a capillary one-way lower stop valve arranged below the condensing pipe body.

[0008] The radial dimension of the condensing pipe body gradually decreases from top to bottom, the capillary one-way lower stop valve is composed of a condensate sample storage area and a condensate sample outlet, the condensate sample storage area is a reverse circular truncated cone structure with a radial dimension gradually decreasing from top to bottom, and the condensate sample outlet is a cylindrical structure with an inner diameter of the order of hundreds of microns.

[0009] The length h1 of the condensing pipe body is 2-5 cm, the width L1 at the inlet is 0.5-1 cm, the length h2 of the condensate sample outlet is 2-3 mm, the inner diameter L2 is of the order of hundreds of microns, and the cutting angle a of the condensate sample storage area ranges from 10 to 60 degrees.

[0010] The peripheral thickness of the cooling trap aluminum block is 0.4-0.6 cm, and gradually thickens from top to bottom on the side close to the thermoelectric cooler Peltier element to fully adhere to the thermoelectric cooler Peltier element.

[0011] The microfluidic chip comprises a microfluidic reaction chamber, a condensate sample inlet, a flushing liquid inlet, and a sample outlet; the on-chip gate graphene field effect transistor biosensor comprises a plurality of groups of graphene field effect transistor devices, the on-chip gate graphene field effect transistor device comprising a source electrode, a drain electrode, and a shared on-chip gate electrode; wherein the graphene biosensing channel of the on-chip gate graphene field effect transistor biosensor is aligned with the microfluidic reaction chamber of the microfluidic chip.

[0012] The method for preparing the graphene biosensing channel of the on-chip gate graphene field effect transistor biosensor comprises: inactivating BSA on the surface of graphene, etching by plasma, and mixing carboxyl-modified polyethylene glycol (PEG) with streptavidin or protein G to modify the surface of the graphene channel by EDC and NHS in the inactivated BSA modified graphene channel; wherein the detection of the novel coronavirus in the respiratory gas condensate of the novel coronavirus infected person can be realized by the directional combination of the novel coronavirus antibody with protein G or the directional modification of the biotin-modified antibody with streptavidin.

[0013] The on-chip gate graphene field effect transistor biosensor is further attached to a PCB board with a signal electrode, and a wire-bonding packaging method is adopted to lead out the signals of the sensor by gold wires or aluminum wires.

[0014] The signal acquisition and control module comprises a microcontroller unit, an analog-to-digital converter, an analog multiplexer, a digital-to-analog converter module, a signal amplifier module, a voltage biasing module, an LCD display module, a voltage regulation module, and a thermoelectric cooling module; wherein the microcontroller unit is connected with the drain electrode of the on-chip gate graphene field effect transistor biosensor through the analog-to-digital converter, the analog multiplexer, and the signal amplifier module, connected with the gate electrode of the on-chip gate graphene field effect transistor biosensor through the digital-to-analog converter module, and connected with the source electrode of the on-chip gate graphene field effect transistor biosensor through the voltage biasing module.

[0015] The microcontroller unit is connected with a temperature sensor through the analog-to-digital converter, the analog multiplexer, and the voltage biasing module, for detecting the surface temperature of the cooling sink aluminum block; the microcontroller unit is also connected with the thermoelectric cooler Peltier element through the analog-to-digital converter, the analog multiplexer, the signal amplifier module, and the voltage regulation module, for maintaining the temperature of the cooling sink aluminum block and the condensate collection tube at 5℃ to -10℃, so as to ensure the collection efficiency of the respiratory gas condensate.

[0016] The microfluidic integrated biosensing system for collecting and detecting respiratory gas condensate provided by the application has the beneficial effects that through the design of the overall structure of the collection tube, the optimized design of the interface of the graphene field effect biosensor, and the design of the capillary one-way stop valve, an integrated respiratory gas condensate detection system that is simple and effective and can realize extremely low target molecule detection is constructed, the system design is simplified, the collection and detection of EBC can be realized at the same time, the risk and work burden faced by the sampling personnel are avoided, and the risk of sample pollution is avoided. BRIEF DESCRIPTION OF DRAWINGS

[0017] Figure 1 is the overall structure schematic diagram of the microfluidic integrated biosensing system for collecting and detecting respiratory gas condensate provided by the application;

[0018] Figure 2 is the partial structure schematic diagram of the microfluidic integrated biosensing system as shown in Figure 1 , wherein the structure of the respiratory gas condensate collector based on thermoelectric cooling is highlighted;

[0019] Figure 3 is the detail enlarged schematic diagram of the condensate collection tube;

[0020] Figure 4 is the assembly schematic diagram of the on-chip gate graphene field effect tube biosensing system based on microfluidic integration;

[0021] Figure 5 is the detection principle diagram of the on-chip gate graphene field effect tube biosensor;

[0022] Figure 6 is the circuit principle structure diagram of the microfluidic integrated biosensing system provided by the application;

[0023] Figure 7 is the detection result of the microfluidic integrated biosensing system for CEA protein detection in application example 1. DETAILED DESCRIPTION

[0024] The application will be further described below in combination with specific embodiments. It should be understood that the following embodiments are only used to illustrate the application and not used to limit the scope of the application.

[0025] In the description of this invention, it should be understood that the terms "center," "longitudinal," "lateral," "length," "width," "thickness," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," "outer," "clockwise," and "counterclockwise," etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are only for the convenience of describing this invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on this invention.

[0026] Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of that feature. In the description of this invention, "a plurality of" means two or more, unless otherwise explicitly specified.

[0027] To address the lack of an integrated detection system for biomolecules in respiratory gas condensates in existing technologies, this invention proposes to combine a thermoelectrically cooled "spindle-shaped" respiratory gas condensate collection tube with a high-sensitivity graphene field-effect transistor biosensor based on on-chip gate (side-gate) control. Through the asymmetrical capillary one-way shut-off valve structure design at both ends of the collection tube, EBC collection and detection can be integrated.

[0028] like Figure 1 The image shows a microfluidic integrated biosensor system for collecting and detecting respiratory gas condensate, provided according to a preferred embodiment of the present invention. It mainly includes: a thermoelectrically cooled respiratory gas condensate collector, a microfluidic integrated on-chip gate graphene field-effect transistor biosensor system, a power supply 8, and a signal acquisition and control module 9. Detailed description follows:

[0029] The thermoelectrically cooled respiratory gas condensate collector includes: a flexible tube 6, a condensate collection tube 1 connected to the flexible tube 6, a cooling trap aluminum block 2 sleeved on the outside of the condensate collection tube 1, a thermoelectric cooler Peltier element 4 and an MCU cooler 7 disposed close to one side of the cooling trap aluminum block 2. A porous filter membrane 10 is provided at the connection between the condensate collection tube 1 and the flexible tube 6. This porous filter membrane is a commercially available, general-purpose filter membrane with a pore size controlled above 10 micrometers, used to filter out saliva and phlegm. The aerosol particle size in the respiratory gas condensate is generally within 5 micrometers. It should be understood that the MCU cooler and the Peltier element 4 can optionally be bonded together with thermally conductive adhesive or thermally conductive tape.

[0030] The microfluidic integrated on-chip gate graphene field effect transistor biosensor system comprises a microfluidic chip 3 and an on-chip gate graphene field effect transistor biosensor 5 assembled by plasma bonding.

[0031] As shown in the drawings, Figure 2 The condensate collection tube 1 comprises a breathing gas introduction tube 11, a condenser tube body 12 connected with the breathing gas introduction tube 11, a capillary one-way upper stop valve 13 arranged above the condenser tube body 12, and a capillary one-way lower stop valve 14 arranged below the condenser tube body 12. The breathing gas introduction tube 11 is connected with one end 61 of the hose 6 through an interface, and the other end 62 of the hose 6 can be directly used for collecting the breathing gas of the patient (as shown in the drawings) or connected with the syringe 20 after the breathing gas collection is completed, so that the remaining condensate in the condensate collection tube 1 is injected into the biosensor 5 through the syringe 20 (as shown in the drawings). Figure 1 The condensate collection tube 1 comprises a breathing gas introduction tube 11, a condenser tube body 12 connected with the breathing gas introduction tube 11, a capillary one-way upper stop valve 13 arranged above the condenser tube body 12, and a capillary one-way lower stop valve 14 arranged below the condenser tube body 12. The breathing gas introduction tube 11 is connected with one end 61 of the hose 6 through an interface, and the other end 62 of the hose 6 can be directly used for collecting the breathing gas of the patient (as shown in the drawings) or connected with the syringe 20 after the breathing gas collection is completed, so that the remaining condensate in the condensate collection tube 1 is injected into the biosensor 5 through the syringe 20 (as shown in the drawings). Figure 2 The condensate collection tube 1 comprises a breathing gas introduction tube 11, a condenser tube body 12 connected with the breathing gas introduction tube 11, a capillary one-way upper stop valve 13 arranged above the condenser tube body 12, and a capillary one-way lower stop valve 14 arranged below the condenser tube body 12. The breathing gas introduction tube 11 is connected with one end 61 of the hose 6 through an interface, and the other end 62 of the hose 6 can be directly used for collecting the breathing gas of the patient (as shown in the drawings) or connected with the syringe 20 after the breathing gas collection is completed, so that the remaining condensate in the condensate collection tube 1 is injected into the biosensor 5 through the syringe 20 (as shown in the drawings).

[0032] The condensate collection tube 1 is preferably made of polytetrafluoroethylene (PTFE) or hydrophobic treated polycarbonate (PC) or polypropylene (PP) material, which has the beneficial effect of improving the collection efficiency of the condensate droplets formed on the surface of the breathing gas. The capillary one-way upper stop valve 13 arranged above the condenser tube body 12 and the capillary one-way lower stop valve 14 arranged below the condenser tube body 12 have the beneficial effect of effectively reducing the backflow of the breathing gas from the condenser tube body 12 to the breathing gas introduction tube 11 and effectively reducing the backflow of the condensate from the biosensor 5 to the condenser tube body 12.

[0033] As shown in the drawings, Figure 3 The radial dimension of the condenser tube body 12 gradually decreases from top to bottom, and the capillary one-way lower stop valve 14 is composed of a condensate sample storage area 122 and a condensate sample outlet 123. The condensate sample storage area 122 adopts a rounded table structure design, and the size of the condenser tube gradually decreases from the upper end of the condenser tube to the lower end of the condenser tube. The condensate sample outlet 123 is a cylindrical area with an inner diameter of about 100 microns. The condensate droplets D formed on the surfaces of the condenser tube walls 124 and 125 will flow into the condensate sample storage area 122 along the tube walls. Due to the gradual reduction of the overall structure size of the condenser tube body 12, the condensate sample storage area 122 and the condensate sample outlet 123, a capillary one-way lower stop valve 14 is formed. Due to the capillary force, the condensate Y temporarily stored in the condensate sample storage area 122 will stay in the condensate sample storage area 122 without external force and will finally flow out from the condensate sample outlet 123. The condensate sample outlet 123 is connected to the microfluidic integrated on-chip gate graphene field effect transistor biosensor system by being nested with the hose 24.

[0034] According to the preferred embodiment, the length h1 of the condensate tube body 12 is 2-5 cm, the width L1 at the inlet of the condensate tube body 12 is 0.5-1 cm, the length h2 of the condensate sample outlet 123 is 2-3 mm, the inner diameter L2 of the condensate sample outlet 123 is in the order of hundreds of microns, and the cut angle a of the condensate sample storage area 122 ranges from 10 to 60 degrees.

[0035] Back to Figure 2 The peripheral thickness of the cooling sink aluminum block 2 matched with the condensate collection tube 1 is 0.4-0.6 cm, and a circular hole 21 is provided at the lower end of the hollow aluminum cylinder to facilitate the outflow of the condensate. The left area 22 of the cooling sink aluminum block gradually thickens from top to bottom, and the right area 23 maintains a uniform thickness from top to bottom. The purpose of this design structure is to maximize the contact area between the cooling sink aluminum block 2 and the Peltier element 4 of the thermoelectric cooler on the one hand, and to ensure sufficient contact between the cooling sink aluminum block 2 and the condensate collection tube 1 on the other hand, thereby ensuring the collection efficiency of the condensate collection tube 1.

[0036] As Figure 4 shown, a side-gate graphene field effect transistor biosensor system based on microfluidic integration is provided according to the present application, which comprises a microfluidic chip 3 and a side-gate graphene field effect transistor biosensor 5 assembled together by plasma bonding. The microfluidic chip 3 comprises a microfluidic reaction chamber 31, a condensate sample inlet 32, a flushing liquid inlet 33, and a sample outlet 34. The microfluidic reaction chamber 31 can be rectangular, used for driving the detection sample on the surface of the biosensor 5 and cleaning the surface of the biosensor 5. The side-gate graphene field effect transistor biosensor 5 has a graphene biosensing channel 51 and four groups of graphene field effect transistor devices, specifically including a source electrode 52, a drain electrode 53, and a shared side-gate 54. It should be understood that the graphene field effect transistor devices are not limited to four groups, which are only used as an example but not as a limitation. The side-gate graphene field effect transistor biosensor 5 is prepared by preparing a gold electrode, transferring graphene, and surface modification on a silicon oxide substrate.

[0037] The microfluidic chip 3 and the side-gate graphene field effect transistor biosensor 5 are Figure 4 integrated together by a plasma bonding process, wherein the graphene biosensing channel 51 of the graphene field effect transistor biosensor 5 is aligned with the microfluidic reaction chamber 31 of the microfluidic chip 3. The integrated design of the side-gate graphene biosensor and the microfluidic chip provided according to the present application has the beneficial effect of facilitating the integration of the graphene biosensor and the construction of the peripheral system.

[0038] As Figure 4As shown below, the on-chip gate graphene field effect transistor biosensor 5 is further attached to a PCB board 26 with a signal electrode 25, the signal electrode 25 is connected with a signal acquisition and control circuit of the transistor, and a wire-bonding packaging mode is adopted to lead out the signal of the sensor through gold wires or aluminum wires. The beneficial effect is that through the packaging mode and the design of the signal acquisition system, the detection signal of the on-chip gate graphene field effect transistor biosensor 5 can be stably output.

[0039] Taking the detection of respiratory gas condensate of a COVID-19 infected person as an example, the construction and detection principle of the on-chip gate graphene field effect transistor biosensor 5 provided by the application are as follows:

[0040] As shown in Figure 5 , first, the bovine serum albumin (BSA) on the graphene surface is thermally inactivated (optionally, 60-90℃, thermal inactivation treatment for 1-10min), etched by plasma, covalently combined through amino and carboxyl groups, mixed with streptavidin or protein G through carboxyl modified polyethylene glycol (PEG) and EDC and NHS in the inactivated BSA modified graphene channel, and combined with protein G or biotin modified antibody and streptavidin, so as to realize the detection of COVID-19 in the respiratory gas condensate of a COVID-19 infected person.

[0041] The beneficial effect of the mixed modification based on polyethylene glycol is that the mixed modification of polyethylene glycol on the surface of the biosensor can increase the Debye length of the biosensing interface, which can effectively increase the response of the graphene field effect transistor biosensor in high salt ion samples. The design of the directional modification based on streptavidin or protein G can increase the efficiency and activity of the antibody modification of the biosensor interface, thereby effectively improving the detection sensitivity and stability of the graphene field effect transistor biosensor.

[0042] As shown in Figure 6 , the signal acquisition and control module 9 of the application includes a microcontroller unit (MCU), an analog-to-digital converter (ADC), an analog multiplexer (Multiplexer, MUX), a digital-to-analog converter module (DAC), a signal amplifier module, a voltage bias module, an LCD display module, a voltage regulation module and a thermoelectric cooling module.

[0043] The microcontroller unit (MCU) is connected to the drain electrode 53 of the on-chip gate graphene field effect transistor biosensor 5 through an analog-to-digital converter (ADC), an analog multiplexer (MUX) and a signal amplifier module, connected to the gate electrode 54 of the on-chip gate graphene field effect transistor biosensor 5 through a digital-to-analog converter module (DAC), and connected to the source electrode 52 of the on-chip gate graphene field effect transistor biosensor 5 through a voltage bias module.

[0044] The DAC, the voltage bias module and the small signal amplifier can control the voltage applied to the source and gate of the graphene field effect transistor biosensor, the output current of the graphene field effect transistor biosensor is returned to the microcontroller unit (MCU) through the amplifier and the analog-to-digital converter (ADC) circuit, data analysis is performed in the microcontroller unit (MCU), and finally the detection concentration of the target molecule is displayed through an external PC terminal display module or other display modules.

[0045] According to the application, the microcontroller unit (MCU) is also connected to the temperature sensor 27 through an analog-to-digital converter (ADC), an analog multiplexer (MUX) and a voltage bias module. The temperature sensor 27 can be a platinum resistance temperature sensor or a thermocouple temperature sensor, and is used to detect the surface temperature of the cooling sink aluminum block 2. The microcontroller unit (MCU) is also connected to the thermoelectric cooler Peltier element 4 through an analog-to-digital converter (ADC), an analog multiplexer (MUX), a signal amplifier module and a voltage regulation module. Through the design of software and hardware, the temperature of the cooling sink aluminum block 2 and the condensate collection tube 1 is maintained at 5℃ to -10℃, so as to ensure the collection efficiency of the respiratory gas condensate.

[0046] According to the application, the microfluidic integrated biosensing system is integrated with the detection process of the biomarker in the respiratory gas condensate as follows:

[0047] First, the condensate collection tube 1 is inserted into the cooling sink aluminum block 2, the lower end of the condensate collection tube 1 is connected to the biosensing system through a luer joint or a hose, and the collection of the respiratory gas condensate and the integration of the biosensor are completed.

[0048] Then, the collection of respiratory gas condensate is carried out, the patient exhales through the hose 6, when the exhaled gas enters the condensate collection tube 1, due to the low temperature of the condensate collection tube 1, the exhaled gas is condensed in the condensate collection tube 1 to form condensate, when enough condensate is accumulated in the condensate collection tube 1, the condensate will automatically flow into the biosensor chamber through the capillary one-way stop valve 14 through the condensate sample outlet 123, if the amount of condensate is small, after the completion of the condensate collection, the exhaled gas condensate can also be injected into the microfluidic chip 3 and the surface of the on-chip gate graphene field effect tube biosensor 5 through the syringe 10;

[0049] Finally, the biosensor signal is collected through the matched signal acquisition and control module 9, so that the detection of biomarkers in respiratory gas condensate is realized.

[0050] Application Example 1

[0051] The microfluidic integrated biosensor system provided by the above embodiment is applied to the detection of CEA protein, and the detection result is as shown in the following table: Figure 7 The sensitivity is as high as 10 pg / mL.

[0052] The above is only a preferred embodiment of the present application, and is not intended to limit the scope of the present application. The above embodiment of the present application can be variously changed. Any simple, equivalent changes and modifications made according to the content of the claims and the description of the present application fall within the scope of the claims of the present application. The present application does not describe all conventional technical contents.

Claims

1. A microfluidic integrated biosensing system for integrated collection and detection of respiratory gas condensate, characterized in that, Comprise: Breathing gas condensate collector based on thermoelectric cooling, comprising: a hose (6), a condensate collection tube (1) connected with the hose (6), a cooling sink aluminum block (2) sleeved outside the condensate collection tube (1), a thermoelectric cooler Peltier element (4) arranged close to one side of the cooling sink aluminum block (2), and an MCU cooler (7); Microfluidic integrated on-chip gate graphene field effect transistor biosensor system, comprising: a microfluidic chip (3) and an on-chip gate graphene field effect transistor biosensor (5) assembled by plasma bonding, the on-chip gate graphene field effect transistor biosensor (5) having a graphene biosensing channel (51) modified with streptavidin or protein G capable of binding with coronavirus antibodies; the preparation method of the graphene biosensing channel comprises: inactivating BSA on the surface of graphene, etching by plasma, and mixing carboxyl modified polyethylene glycol (PEG) with streptavidin or protein G to modify the surface of the graphene channel by EDC and NHS; wherein the coronavirus antibodies are combined with protein G or biotin modified antibodies are combined with streptavidin, so as to realize the detection of coronavirus in the respiratory gas condensate of coronavirus infected persons; Power supply (8) and signal acquisition and control module (9); The condensate collection tube (1) comprises: a breathing gas inlet pipe (11), a condensing pipe body (12) connected with the breathing gas inlet pipe (11), a capillary one-way upper stop valve (13) arranged above the condensing pipe body (12), and a capillary one-way lower stop valve (14) arranged below the condensing pipe body (12); the radial dimension of the condensing pipe body (12) gradually decreases from top to bottom, the capillary one-way lower stop valve (14) is composed of a condensate sample storage area (122) and a condensate sample outlet (123), the condensate sample storage area (122) is a reverse circular table structure with a radial dimension gradually decreasing from top to bottom, and the condensate sample outlet (123) is a cylindrical structure with an inner diameter of the order of hundreds of microns; the length h1 of the condensing pipe body (12) is 2-5 cm, the width L1 at the inlet is 0.5-1 cm, the length h2 of the condensate sample outlet (123) is 2-3 mm, the inner diameter L2 is of the order of hundreds of microns, and the cutting angle α of the condensate sample storage area (122) ranges from 10 to 60 degrees; Wherein, the exhaled gas enters the condensate collection tube (1) through the hose (6) and condenses, the formed condensate flows into the microfluidic chip (3) and the surface of the on-chip gate graphene field effect transistor biosensor (5), and the biosensor signal is collected by the signal acquisition and control module (9), so as to realize the detection of biomarkers in the breathing gas condensate.

2. The microfluidic integrated biosensing system of claim 1, wherein, The peripheral thickness of the cooling sink aluminum block (2) is 0.4-0.6 cm, and gradually thickens from top to bottom on the side close to the thermoelectric cooler Peltier element (4) to fully adhere to the thermoelectric cooler Peltier element (4).

3. The microfluidic integrated biosensing system of claim 1, wherein, The microfluidic chip (3) comprises a microfluidic reaction chamber (31), a condensate sample inlet (32), a flushing liquid inlet (33), and a sample outlet (34); the on-chip gate graphene field effect transistor biosensor (5) comprises a plurality of groups of graphene field effect transistor devices, and the graphene field effect transistor devices further comprise a source electrode (52), a drain electrode (53), and a shared on-chip gate electrode (54); wherein the graphene biosensing channel (51) of the graphene field effect transistor biosensor (5) is aligned with the microfluidic reaction chamber (31) of the microfluidic chip (3).

4. The microfluidic integrated biosensing system of claim 3, wherein, The on-chip gate graphene field effect transistor biosensor (5) is further attached to a PCB board (26) with a signal electrode (25), and adopts a wire bonding packaging mode to output the sensor signal through gold wires or aluminum wires.

5. The microfluidic integrated biosensing system of claim 3, wherein, The signal acquisition and control module (9) comprises a microcontroller unit, an analog-to-digital converter, an analog multiplexer, a digital-to-analog converter module, a signal amplifier module, a voltage biasing module, an LCD display module, a voltage regulating module, and a thermoelectric cooling module, wherein the microcontroller unit is connected to the drain electrode (43) of the on-chip gate graphene field effect transistor biosensor (5) through the analog-to-digital converter, the analog multiplexer, and the signal amplifier module, connected to the gate electrode (44) of the on-chip gate graphene field effect transistor biosensor (5) through the digital-to-analog converter module, and connected to the source electrode (42) of the on-chip gate graphene field effect transistor biosensor (5) through the voltage biasing module.

6. The microfluidic integrated biosensing system of claim 5, wherein, The microcontroller unit is connected to a temperature sensor (27) through the analog-to-digital converter, the analog multiplexer, and the voltage biasing module, for detecting the surface temperature of the cooling sink aluminum block (2); the microcontroller unit is also connected to the thermoelectric cooler Peltier element (4) through the analog-to-digital converter, the analog multiplexer, the signal amplifier module, and the voltage regulating module, to maintain the temperature of the cooling sink aluminum block (2) and the condensate collection tube (1) at 5℃~-10℃, so as to ensure the collection efficiency of the respiratory gas condensate.

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