Optical fiber DNA sensor prepared by combining molybdenum disulfide material and detection method

Abstract

The invention provides an optical fiber DNA sensor prepared by combining a molybdenum disulfide material and a detection method, a micro-fluidic chip with a tip coupled bubble structure is provided, the micro-fluidic chip comprises an elastic material chip and a glass sheet which are bonded, the plane of the glass sheet covers the plane projection part of the elastic material chip, and the plane projection part of the elastic material chip covers the plane projection part of the glass sheet. One plane of the elastic material chip is used as a bonding plane to be bonded with the glass sheet, a groove penetrating through the length direction of the elastic material chip is formed in the bonding plane, and the groove is used for placing the optical fiber biosensor in the length direction and is filled with liquid acting on the optical fiber biosensor; the groove comprises a liquid channel, a first extension area, a second extension area and a plurality of tip coupling bubble structures, the first extension area and the second extension area are located on the two sides of the liquid channel in the length direction respectively, and the tip coupling bubble structures are located on one side of the liquid channel in the width direction.

Description

Technical Field

[0001] The present application relates to the technical field of optical biosensors, and in particular to an optical fiber DNA sensor prepared in combination with molybdenum disulfide material and a detection method. Background Art

[0002] Molybdenum disulfide (MoS2) fiber biosensors utilize optical fiber technology to detect target biomolecules. They are widely used in biomedical applications such as disease detection, genetic analysis, and drug screening. Compared to traditional polymerase chain reaction (PCR) and electrophoresis-based detection methods, MoS2 fiber biosensors offer high sensitivity, low cost, rapid response, and parallel detection capabilities. When target DNA (deoxyribonucleic acid) molecules present in a liquid sample complement the DNA probes modified on the MoS2 fiber biosensor, a binding reaction occurs on the sensor surface. By monitoring the changes in the optical signal caused by this reaction, the presence and concentration of the target DNA can be quantitatively analyzed in real time, enabling the detection of gene mutations, pathogens, or cancer-related genes. This approach holds important implications for early disease diagnosis, genetic disease detection, and cancer gene screening. Various approaches have been developed to enhance the sensitivity of MoS2 fiber DNA sensors, focusing on biosignal amplification, structural material improvements, and optimized contact properties.

[0003] For biosignal amplification, well-controlled polymers are produced by coupling alkyl halide free radical initiators with copper-based redox catalysts. When the film exhibits opacity, poly(hydroxyethyl methacrylate) polyhema specifically binds to the complementary sequence; when a non-complementary sequence is applied, the film remains transparent. This method can be used to detect single-base mismatched DNA sequences in liquid samples. Another type of signal amplification is the use of nanocatalysts to drive chemical reactions, which further enhances the detection results and thus improves the detection sensitivity. Platinum-modified gold nanorods (AU / PT NPs) can be used as nanocatalysts for ultrasensitive colorimetric assays for in vitro diagnostics, capable of producing color signals several orders of magnitude higher.

[0004] In terms of structural material improvements, high-performance optical fiber sensors with a hybrid structure of two-dimensional nanomaterials (such as BP-WS, a composite material composed of black phosphorus (BP) and tungsten disulfide (WS2)) are used to detect DNA hybridization. DNA molecules aggregate on the BP-WS surface, thereby improving the sensitivity of detecting target DNA molecules, which is 1.46 times more sensitive than traditional sensors without a hybrid structure. Alternatively, hollow tube lattice fiber (HC-TLF) can be considered as a label-free biosensor for detecting target DNA molecules. By comparing the optical fiber transmission spectra before and after flow, the final formation of the binding layer can be determined, thereby determining whether the target DNA sequence is in the solution.

[0005] In the study of optimizing contact effects, molecular recognition in biosensing is controlled by the collision of interacting molecules and the subsequent binding reaction. Methods that can increase the probability of contact between the sensor surface and the target analyte can be used to improve the sensitivity of surface-based sensors. By applying current to the electrode array in a microfluidic device, a vortex-like microflow pattern is generated through the electrodes, which concentrates the dispersed proteins on the substrate surface and drives the mixing of the fluid. Using surface-immobilized disease-specific antigens and fluorescent secondary antibodies as capture probes, the fluorescence detection speed is increased tenfold compared to traditional microfluidic devices. Alternatively, by utilizing a superhydrophobic surface, the target detection object can be confined to a defined sensing area. In the prepared micropillar array, its surface has extremely high hydrophobicity, making the contact angle of the liquid much greater than 90° (>150°), which enables the target molecules to be stably concentrated in the stably shrinking quasi-spherical droplets.

[0006] The molecular recognition process in biosensing is governed by the collision of interacting molecules and the subsequent binding reaction. While the binding reaction relies on the binding affinity between the two molecules, which is determined by the molecular structure and reaction mechanism, the occurrence of collisions is affected by the diffusion of the target molecule in the solution to the surface-immobilized probe. In practical applications, the binding substrate is usually incubated with the target molecule solution. For example, each incubation step in the enzyme-linked immunosorbent assay (ELISA) requires a long time. When the concentration of the analyte is diluted, it usually takes a long time for the binding to be transduced into a detectable signal in the diffusion-based incubation. This shortcoming greatly restricts the efficiency and performance of sensing detection, especially the sensitivity. Summary of the Invention

[0007] In view of this, the purpose of this application is to provide at least a fiber optic DNA sensor and detection method prepared in combination with molybdenum disulfide material, by designing a microfluidic chip containing a tip-coupled bubble structure, and combining the microfluidic chip with a piezoelectric transducer to design a fiber optic DNA sensor, and increasing the contact probability between the sensor and the molecule to be detected by the tip-coupled bubble structure and ultrasonic enhancement, thereby solving the technical problem of low detection sensitivity in the existing technology and achieving the technical effect of increasing detection sensitivity.

[0008] This application mainly includes the following aspects:

[0009] In a first aspect, an embodiment of the present application provides a microfluidic chip with a tip-coupled bubble structure, the microfluidic chip comprising a bonded elastic material chip and a glass sheet, wherein the plane of the glass sheet covers the planar projection portion of the elastic material chip, a plane of the elastic material chip serves as a bonding plane for bonding with the glass sheet, a groove is provided on the bonding plane running through the length direction of the elastic material chip, the groove is used to place an optical fiber biosensor in the length direction and to be filled with liquid acting on the optical fiber biosensor, the groove comprises a liquid channel, a first extension area, a second extension area and a plurality of tip-coupled bubble structures, the groove comprises a liquid channel, a first extension area, a second extension area and a plurality of A tip-coupled bubble structure, the first extension area and the second extension area are respectively located on both sides of the liquid channel in the length direction, the multiple tip-coupled bubble structures are located on one side of the width direction of the liquid channel, the width of the liquid channel in the width direction is greater than the width of the extension area in the width direction, the depth of the liquid channel in the depth direction is less than the depth of the extension area in the depth direction, the depth of the tip-coupled bubble structure in the depth direction is equal to the depth of the liquid channel in the depth direction, the tip-coupled bubble structure is a fan-shaped structure, one side of the fan-shaped structure is connected to the liquid channel, and the other side of the fan-shaped structure does not contact the liquid channel.

[0010] Optionally, the first extension area includes a first optical fiber connection port and a first liquid flow port, and the first extension area includes a second optical fiber connection port and a second liquid flow port, wherein the first optical fiber connection port and the second optical fiber connection port are located on both sides of the liquid channel in the length direction, the first liquid flow port is located on one side of the first optical fiber connection port in the width direction, and the second liquid flow port is located on one side of the second optical fiber connection port in the width direction, and the specifications of the optical fiber connection ports are set according to the cross-section of the optical fiber biosensor so that the first optical fiber connection port, the liquid channel and the second optical fiber connection port are used to place the optical fiber biosensor in the length direction; and / or, the material of the elastic material chip includes polydimethylsiloxane, a curing agent and a surfactant.

[0011] In the second aspect, an embodiment of the present application also provides a method for preparing a microfluidic chip, the preparation method comprising: placing a silicon wafer master mold in a fume hood, and dripping trimethylchlorosilane onto the silicon wafer master mold for sealing treatment for a first preset time period, wherein the surface of the silicon wafer master mold is provided with at least one channel for preparing the elastic material chip; pouring a mixed solution of the elastic material chip material onto the surface of the silicon wafer master mold for heating and curing, and peeling the cooled elastic material chip off the silicon wafer master mold; making a groove on the bonding plane of the elastic material chip by a punch; performing a bonding operation on the bonding plane of the elastic material chip and a glass sheet to prepare the microfluidic chip; wherein the microfluidic chip refers to the microfluidic chip as described in the first aspect or any possible embodiment of the first aspect.

[0012] Optionally, the silicon wafer master mold is prepared in the following manner: after cleaning the silicon wafer, the silicon wafer is placed on a heating plate for baking; the baked silicon wafer is fixed on the operating table of a coating machine, and photoresist is coated on the surface of the silicon wafer; after placing the coated silicon wafer on a heating plate for baking, a mask is placed on the photoresist on the surface of the silicon wafer for exposure, and the mask includes at least one channel pattern corresponding to the elastic material chip; after placing the exposed silicon wafer on a heating plate for baking, the silicon wafer is developed and fixed to generate at least one channel for preparing the elastic material chip on the surface of the silicon wafer; the silicon wafer is hardened to prepare the silicon wafer master mold.

[0013] Optionally, the bonding operation is performed on the bonding plane of the elastic material chip and the glass sheet in the following manner: the glass sheet is cleaned with anhydrous ethanol and deionized water, and then the cleaned glass sheet is blown dry with compressed nitrogen; the elastic material chip and the glass sheet are surface-treated with a plasma bonding machine, and after the surface treatment, the bonding plane of the elastic material chip and the surface of the glass sheet are adhered to complete the bonding.

[0014] In a third aspect, an embodiment of the present application further provides an optical fiber DNA sensor prepared in combination with molybdenum disulfide material, the sensor comprising: a microfluidic chip as described in the first aspect or any possible embodiment of the first aspect, wherein a circular hole is provided on a glass sheet of the microfluidic chip; a piezoelectric transducer, wherein a vibrating plate of the piezoelectric transducer is provided corresponding to the circular hole so as to expose the vibrating plate through the circular hole; a first drainage device and a second drainage device, wherein the first drainage device is connected to the first liquid flow port, and the second drainage device is connected to the second liquid flow port, so as to control the flow of liquid into and out of the liquid channel through the first drainage device and the second drainage device; an optical fiber DNA sensor fixed to an area on the glass sheet other than the piezoelectric transducer; wherein the bonding plane of the elastic material chip of the microfluidic chip is placed on the glass sheet for bonding, so that the groove of the elastic material chip is located above the optical fiber DNA sensor, and the sensing area of ​​the optical fiber DNA sensor is located in the liquid channel, and the first optical fiber communication port and the second optical fiber communication port are respectively sealed after the optical fiber DNA sensor is placed.

[0015] Optionally, the fiber-optic DNA sensor is prepared by: after cleaning the surface of the fiber-optic DNA sensor to be treated with deionized water, the fiber-optic DNA sensor is immersed in a piranha solution to remove contaminants and activate hydroxyl groups on the surface of the sensing area of ​​the fiber-optic DNA sensor, and then the fiber-optic DNA sensor is washed with deionized water; after fixing the fiber-optic DNA sensor on the glass slide, the sensing area is coated with molybdenum disulfide a preset number of times to prepare the fiber-optic DNA sensor, wherein the molybdenum disulfide coating is achieved by coating the sensing area with a molybdenum disulfide aqueous solution and then placing the glass slide on a temperature control console for heating and evaporation.

[0016] In a fourth aspect, an embodiment of the present application further provides a DNA detection method, which is applied to the optical fiber DNA sensor as described in the third aspect or any possible embodiment of the third aspect, wherein the method includes: preparing a DNA probe complementary to the DNA to be tested on the sensing area of ​​the optical fiber DNA sensor in the optical fiber DNA sensor; injecting a test solution containing the DNA to be tested into the liquid channel through the first drainage device and the second drainage device, so that the DNA to be tested and the DNA probe achieve DNA complementarity in the sensing area; collecting a spectral curve of the optical fiber DNA sensor after DNA complementation is achieved, the spectral curve being used to reflect the intensity of light corresponding to different wavelengths; and determining a target DNA concentration corresponding to the target wavelength according to the target wavelength corresponding to the characteristic peak on the spectral curve, so as to use the target DNA concentration as the DNA concentration to be tested of the test solution.

[0017] Optionally, the DNA probe is prepared in the following manner: a polylysine solution is injected into the liquid channel through the first drainage device and the second drainage device, and after the polylysine solution infiltrates the optical fiber DNA sensor for a second preset time period, the optical fiber DNA sensor is cleaned with deionized water; a probe solution containing a DNA probe is injected into the liquid channel through the first drainage device and the second drainage device, and the probe solution is controlled to infiltrate the optical fiber DNA sensor for a third preset time period so that the DNA probe adheres to the optical fiber DNA sensor, and the optical fiber DNA sensor is cleaned with TE buffer to prepare the DNA probe.

[0018] Optionally, the target DNA concentration corresponding to the target wavelength is determined in the following manner: in a preliminary testing stage, spectral curves corresponding to different preset DNA concentrations are obtained to determine the characteristic wavelengths of the characteristic peaks corresponding to different preset DNA concentrations; data fitting is performed according to the characteristic wavelengths corresponding to each preset DNA concentration to determine a fitting function for reflecting the relationship between DNA concentration and wavelength; and the target DNA concentration corresponding to the target wavelength is determined by the fitting function.

[0019] An embodiment of the present application provides an optical fiber DNA sensor and detection method prepared in combination with molybdenum disulfide material. The microfluidic chip includes a bonded elastic material chip and a glass sheet, wherein the plane of the glass sheet covers the planar projection portion of the elastic material chip, and a plane of the elastic material chip is used as a bonding plane for bonding with the glass sheet. A groove is provided on the bonding plane that runs through the length direction of the elastic material chip. The groove is used to place an optical fiber biosensor in the length direction and fill it with liquid that acts on the optical fiber biosensor. The groove includes a liquid channel, a first extension area, a second extension area, and a plurality of tip-coupled bubble structures. The first extension area The extension region and the second extension region are respectively located on both sides of the liquid channel in the length direction, and the multiple tip-coupled bubble structures are located on one side of the width direction of the liquid channel. The width of the liquid channel in the width direction is greater than the width of the extension region in the width direction, the depth of the liquid channel in the depth direction is less than the depth of the extension region in the depth direction, the depth of the tip-coupled bubble structure in the depth direction is equal to the depth of the liquid channel in the depth direction, and the tip-coupled bubble structure is a fan-shaped structure, one side of the fan-shaped structure is connected to the liquid channel, and the other side of the fan-shaped structure does not contact the liquid channel. By designing a microfluidic chip containing a tip-coupled bubble structure and combining the microfluidic chip with a piezoelectric transducer to design an optical fiber DNA sensor, the contact probability between the sensor and the molecule to be detected is increased through the tip-coupled bubble structure and ultrasonic enhancement, thereby solving the technical problem of low detection sensitivity in the prior art and achieving the technical effect of increasing detection sensitivity.

[0020] In order to make the above-mentioned objects, features and advantages of the present application more obvious and easy to understand, preferred embodiments are given below and described in detail with reference to the accompanying drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the following is a brief introduction to the drawings required for use in the embodiments. It should be understood that the following drawings only show certain embodiments of the present application and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other relevant drawings can be obtained based on these drawings without creative work.

[0022] Figure 1 A schematic diagram of a microfluidic chip with a tip-coupled bubble structure provided in an embodiment of the present application is shown.

[0023] Figure 2 A schematic diagram of an elastic material chip provided in an embodiment of the present application is shown.

[0024] Figure 3A schematic diagram showing the steps of the method for preparing a microfluidic chip provided in an embodiment of the present application.

[0025] Figure 4 A schematic diagram showing the steps of the method for preparing a silicon wafer master mold provided in an embodiment of the present application.

[0026] Figure 5 The schematic diagram of the optical fiber DNA sensor provided in the embodiment of the present application is shown. Figure 1 .

[0027] Figure 6 The schematic diagram of the optical fiber DNA sensor provided in the embodiment of the present application is shown. Figure 2 .

[0028] Figure 7 A schematic diagram showing the concentration analysis principle of the optical fiber DNA sensor provided in an embodiment of the present application is shown.

[0029] Figure 8 A schematic diagram of the spectral curve of repeated tests provided in the embodiments of the present application is shown.

[0030] Figure 9 A schematic diagram showing the spectral curves corresponding to different DNA concentrations provided in the examples of the present application.

[0031] Figure 10 A schematic diagram showing the fitting function of the relationship between DNA concentration and wavelength provided in the examples of the present application is shown.

[0032] Figure 11 A schematic diagram showing characteristic wavelengths corresponding to the presence or absence of ultrasonic driving provided in an embodiment of the present application is shown. DETAILED DESCRIPTION

[0033] In order to make the purpose, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. It should be understood that the drawings in the present application only serve the purpose of illustration and description and are not used to limit the scope of protection of the present application. In addition, it should be understood that the schematic drawings are not drawn to scale. The flowcharts used in this application illustrate the operations implemented according to some embodiments of the present application. It should be understood that the operations of the flowcharts can be implemented out of sequence, and steps without logical context can be reversed or implemented simultaneously. In addition, those skilled in the art, under the guidance of the contents of this application, can add one or more other operations to the flowchart, or remove one or more operations from the flowchart.

[0034] In addition, the described embodiments are only a part of the embodiments of the present application, rather than all of the embodiments. The components of the embodiments of the present application generally described and shown in the drawings here can be arranged and designed in various configurations. Therefore, the following detailed description of the embodiments of the present application provided in the drawings is not intended to limit the scope of the claimed application, but merely represents selected embodiments of the present application. Based on the embodiments of the present application, all other embodiments obtained by those skilled in the art without making creative work are within the scope of protection of the present application.

[0035] In existing technologies, the molecular recognition process in biosensing is governed by the collision of interacting molecules and the subsequent binding reaction. While the binding reaction relies on the binding affinity between the two molecules, determined by their molecular structure and reaction mechanism, the occurrence of collisions is affected by the diffusion of the target molecule from the solution to the surface-immobilized probe. When the analyte concentration is dilute, binding typically takes a long time to transduce into a detectable signal during diffusion-based incubation. This limitation significantly limits the efficiency and performance of sensing detection, particularly sensitivity.

[0036] Based on this, the present invention provides an optical fiber DNA sensor and detection method prepared in combination with molybdenum disulfide material. By designing a microfluidic chip containing a tip-coupled bubble structure and combining the microfluidic chip with a piezoelectric transducer to design an optical fiber DNA sensor, the tip-coupled bubble structure and ultrasonic enhancement are used to increase the contact probability between the sensor and the molecule to be detected. This solves the technical problem of low detection sensitivity in the prior art and achieves the technical effect of increasing detection sensitivity. The details are as follows:

[0037] See also Figure 1 , Figure 1 Schematic diagram of a microfluidic chip with a tip-coupled bubble structure provided in an embodiment of the present application. Figure 1 As shown, the microfluidic chip provided in an embodiment of the present application includes a bonded elastic material chip 101 and a glass sheet 102. The plane of the glass sheet covers the planar projection of the elastic material chip, and a plane of the elastic material chip serves as a bonding plane for bonding to the glass sheet. The bonding plane is provided with a groove running through the length direction of the elastic material chip.

[0038] Furthermore, the elastic material chip and the glass sheet can both be understood as rectangular parallelepipeds, the bonding plane of the elastic material chip is no larger than the contact surface of the glass sheet, and the contact surface of the glass sheet refers to a plane on the glass sheet that is bonded to the elastic material chip. Furthermore, the length of the elastic material chip in the length direction is greater than the width of the elastic material chip in the width direction. A plane on the elastic material chip that is parallel to the bonding plane is referred to as a non-bonding plane. The length direction of the non-bonding plane is referred to as the direction of the x-axis, the width direction of the non-bonding plane is referred to as the direction of the y-axis, and the direction of the non-bonding plane pointing vertically downward to the bonding plane is referred to as the depth direction, which is referred to as the direction of the z-axis. Furthermore, in order to bond the glass sheet and the bonding plane of the elastic material chip, the plane projection portion of the bonding plane of the elastic material chip projected onto the glass sheet along the vertical direction should be located on the contact surface of the glass sheet, or in other words, the plane projection portion of the bonding plane of the elastic material chip on the contact surface of the glass sheet should be included.

[0039] Specifically, the groove is used to place the optical fiber biosensor in the length direction and fill it with liquid that acts on the optical fiber biosensor. The groove includes a liquid channel A, a first extension area 1, a second extension area 2 and a plurality of tip-coupled bubble structures B. The first extension area and the second extension area are respectively located on both sides of the liquid channel in the length direction. The plurality of tip-coupled bubble structures are located on one side of the width direction of the liquid channel. The width of the liquid channel in the width direction is greater than the width of the extension area in the width direction. The depth of the liquid channel in the depth direction is less than the depth of the extension area in the depth direction, so that the liquid filled in the liquid channel and the optical fiber biosensor can fully act.

[0040] In other words, the total length of the groove in the longitudinal direction is the same as the length of the elastic material chip in the longitudinal direction, so that when the optical fiber biosensor is placed in the groove, it extends through the length of the elastic material chip. Furthermore, the length of the optical fiber biosensor can be greater than the length of the elastic material chip, and both ends of the optical fiber biosensor can be extended from the first extension region and the second extension region.

[0041] See also Figure 2 , Figure 2 This is a schematic diagram of the elastic material chip provided in the embodiment of the present application. Figure 2As shown, the first extension area includes a first optical fiber connection port L1 and a first liquid flow port L3, and the first extension area includes a second optical fiber connection port L2 and a second liquid flow port L4, wherein the first optical fiber connection port and the second optical fiber connection port are located on both sides of the liquid channel in the length direction, the first liquid flow port is located on one side of the first optical fiber connection port in the width direction, and the second liquid flow port is located on one side of the second optical fiber connection port in the width direction, and the specifications of the optical fiber connection ports are set according to the cross-section of the optical fiber biosensor so that the first optical fiber connection port, the liquid channel and the second optical fiber connection port are used to place the optical fiber biosensor in the length direction.

[0042] Exemplarily, the liquid channel, the first optical fiber connection port L1, and the first liquid flow port L3 can be understood as a three-way pipe located on one side of the liquid channel in the longitudinal direction, and the liquid channel, the second optical fiber connection port L2, and the second liquid flow port L4 can be understood as a three-way pipe located on the other side of the liquid channel in the longitudinal direction. The first optical fiber connection port L1 and the second optical fiber connection port L2 are used to place the optical fiber DNA sensor, so that the optical fiber DNA sensor passes through the first optical fiber connection port L1, the liquid channel, and the second optical fiber connection port L2. The first liquid flow port L3 and the second liquid flow port L4 are used to control the flow of liquid into and out of the liquid channel, that is, liquid flows into the liquid channel from one of the first liquid flow port L3 and the second liquid flow port L4, and liquid flows out of the liquid channel from the other of the first liquid flow port L3 and the second liquid flow port L4.

[0043] Specifically, the width of the liquid channel is greater than that of the first and second optical fiber ports. This increases the space in the liquid channel, allowing for sufficient contact between the liquid and the fiber-optic DNA sensor, thereby increasing the sensor's sensitivity. Because the fiber-optic DNA sensor is a tapered fiber, the diameters at both ends of the sensor are greater than the diameter of the sensing area. Consequently, the depths of the first and second optical fiber ports should be greater than the depth of the liquid channel.

[0044] The depth of the tip-coupled bubble structure in the depth direction is equal to the depth of the liquid channel in the depth direction. Furthermore, the tip-coupled bubble structure is a fan-shaped structure, one side of which is connected to the liquid channel, while the other side of the fan-shaped structure does not contact the liquid channel. Furthermore, the vibration of the vibrating plate of the piezoelectric transducer can drive bubbles in the liquid to the tip-coupled bubble structure. Ultrasonic enhancement can be used to move the bubbles away from the sensing area of ​​the fiber-optic DNA sensor, thereby enhancing the interaction efficiency between the liquid and the sensing area of ​​the fiber-optic DNA sensor and improving sensitivity.

[0045] Exemplarily, the width of the first liquid circulation port L3 and the second liquid circulation port L4 is 1.4 millimeters (mm) so that the liquid injected into the liquid channel has a certain buffering force. Since the widest diameter length on the cross section of the optical fiber is about 15 microns (μm), the cross section can be understood as the cross section obtained by cutting the optical fiber through the plane formed by the y-axis and the z-axis. The width of the first optical fiber connection port and the second optical fiber connection port should be greater than the widest diameter length of the optical fiber, and the width of the first optical fiber connection port L1 and the second optical fiber connection port L2 in the width direction is also set to 1.4mm. In order to ensure that the tip-coupled bubble structure can maintain a good elastic effect under ultrasonic drive, and to ensure that the optical fiber can be embedded in the first optical fiber connection port L1 and the second optical fiber connection port L2 during heterogeneous integration, the depth of the liquid channel is 40μm, and the optimal depth of the first optical fiber connection port and the second optical fiber connection port is 300μm. With the above structure, the optical fiber DNA sensor can be properly placed and the liquid sample can be conveniently placed in and out of the liquid channel.

[0046] The liquid channel may be a long straight channel, or may be configured in an arc shape at both ends connected to the first optical fiber communication port L1 and the second optical fiber communication port L2 respectively.

[0047] like Figure 2As shown, the liquid channel is embedded in a tip-coupled bubble structure. The tip-coupled bubble structure has a sector radius of 170 μm and a central angle of 165°. That is, the angle between one side of the sector structure connecting to the liquid channel and the other side of the sector structure is 165°, and the angle between the other side of the sector structure and the liquid channel is 15°. This allows the liquid to quickly pass through the tip-coupled bubble structure, effectively utilizing the liquid contact angle to form a bubble structure at the tip. Specifically, multiple tip-coupled bubble structures are provided along one side of the liquid channel in the width direction, each projecting outward from one side of the liquid channel in the width direction. A first liquid flow port L3 is provided on one side of the first optical fiber connection port L1 in the width direction (y-axis direction), and a second liquid flow port L4 is provided on one side of the second optical fiber connection port L2 in the width direction (y-axis direction). The widths a1 of the first and second liquid flow ports L3 and L4 are equal to the widths a2 of the first and second liquid flow ports L3 and L4 in the width direction. Furthermore, after the liquid enters the liquid channel through one of the first and second liquid flow ports L3 and L4, the vibration of the piezoelectric transducer's vibrating plate tends to attract bubbles within the liquid to the tip-coupled bubble structure, moving the bubbles away from the sensing area of ​​the fiber-optic DNA sensor. The liquid then flows out through the other of the first and second liquid flow ports L3 and L4. Thus, the tip-coupled bubble structure increases the disturbance of the liquid within the liquid channel, thereby increasing the contact between the fiber-optic DNA sensor and the liquid within the liquid channel and enhancing the sensitivity of the fiber-optic DNA sensor.

[0048] The elastic material chip comprises polydimethylsiloxane (PDMS), a curing agent, and a surfactant. In other words, in order to achieve the perturbation of the liquid by the tip-coupled bubble structure, the elastic polydimethylsiloxane material was selected to make the elastic material chip.

[0049] Based on the same application concept, the embodiments of the present application also provide a method for preparing a microfluidic chip corresponding to the microfluidic chip provided in the above embodiments. Since the principle of solving the problem by the microfluidic chip in the embodiments of the present application is similar to the method for preparing the microfluidic chip in the above embodiments of the present application, the implementation of the method can refer to the implementation of the microfluidic chip, and the repeated parts will not be repeated.

[0050] See also Figure 3 , Figure 3 Schematic diagram of the steps of the method for preparing the microfluidic chip provided in the embodiment of the present application. Figure 3 As shown, the method for preparing the microfluidic chip provided in the embodiment of the present application includes the following steps:

[0051] S101: placing a silicon wafer master mold in a fume hood, and dripping trimethylchlorosilane onto the silicon wafer master mold for sealing for a first preset period of time.

[0052] The silicon wafer master mold has at least one channel on its surface for preparing the elastic material chip. That is, the elastic material chip is prepared by pouring a mixed solution of materials for preparing the elastic material chip into the at least one channel on the silicon wafer master mold, with one channel being used to prepare one elastic material chip.

[0053] Specifically, the silicon wafer master mold is prepared in the following manner: after cleaning the silicon wafer, the silicon wafer is placed on a heating plate for baking; the baked silicon wafer is fixed on the operating table of a coating machine, and photoresist is coated on the surface of the silicon wafer; after the coated silicon wafer is placed on a heating plate for baking, a mask is placed on the photoresist on the surface of the silicon wafer for exposure, and the mask includes at least one channel pattern corresponding to the elastic material chip; after the exposed silicon wafer is placed on a heating plate for baking, the silicon wafer is developed and fixed to generate at least one channel for preparing the elastic material chip on the surface of the silicon wafer; the silicon wafer is hardened to prepare the silicon wafer master mold.

[0054] For example, see Figure 4 , Figure 4 Schematic diagram of the steps of the method for preparing a silicon wafer master mold provided in the embodiment of the present application. Figure 4As shown, the first step is silicon wafer pretreatment: using tweezers, a 4-inch single-polish silicon wafer is cleaned alternately with acetone, anhydrous ethanol, and deionized water. A nitrogen gun is then used to blow dry the wafer surface to remove surface dust. Finally, the oven temperature is set to 200°C, and the wafer is placed on a hot plate inside the oven and left to stand for approximately two hours. This baking process accelerates surface oxidation and enhances adhesion between the wafer and the photoresist. The second step is photoresist coating and spin coating: the pretreated wafer is placed on the operating table of a spin coater, which applies negative pressure to the wafer to secure it in place. Then set two periods of the coating stage: the first period rotates at a speed of 500r / min (revolutions per minute) for 30 seconds, and the second period rotates at a speed of 2700r / min for 60 seconds. Start the coating machine to slowly drip the SU8-2025 model photoresist onto the center of the silicon wafer. According to the speed and time of the two periods set above, control the coating outlet of the coating machine to rotate so that the photoresist is coated on the silicon wafer, ensuring that no bubbles are generated to complete the spin coating. The third step is pre-baking: Place the silicon wafer with the photoresist spun on a hot plate and heat bake it at 65°C for 30 minutes, then at 95°C for 30 minutes. After the baking is completed, wipe the back of the silicon wafer (i.e., the side adsorbed by the operating table) with a small amount of acetone solution dipped in clean paper to remove the residual photoresist on the back. The fourth step is exposure: The prepared front of the mask is placed opposite the photoresist-coated side of the silicon wafer. The front of the mask, usually the chrome film or the patterned side, must face the photoresist, while the back of the mask, usually the glass substrate or the non-patterned side, faces the exposure light source. If the front and back are reversed, the pattern will be blurred, distorted, or even completely unrecognizable, resulting in the wafer being scrapped. The mask also includes a marking line pattern, which is located between the individual channel patterns to distinguish them.

[0055] Then, after placing the mask, vacuum suction is used to fix the silicon wafer to the tray of the photolithography machine, and the mask and silicon wafer are aligned. Then, ultraviolet light of a specific wavelength is used to selectively expose the silicon wafer covered with the mask, so that the photosensitive area undergoes a chemical reaction and can be preserved during the development process. The exposure time is 5 seconds. The exposed silicon wafer is removed. The entire photolithography process is performed under yellow light to prevent the denaturation of the photoresist. Contact exposure can be selected as the exposure method. The fifth step is post-baking: In order to accelerate the chemical reaction of the photosensitive area on the surface of the silicon wafer and weaken the standing wave effect between the exposed and non-exposed areas, the silicon wafer needs to be placed on a hot plate for processing after exposure. It is heated at 95°C for an additional 30 minutes to complete the post-baking. The sixth step is development and fixing: Place the exposed silicon wafer into a container filled with developer and shake it slowly for 3 minutes to make the pattern structure appear completely. In the middle, you can use a rubber-tipped dropper to absorb the developer and slowly rinse the graphic area to enhance the development effect. Then use tweezers to transfer the silicon wafer to a container filled with isopropyl tone and soak it for about 2 minutes to remove the residual glue and developer on the surface of the silicon wafer. Finally, use nitrogen to blow dry the surface of the silicon wafer to complete the fixing. The seventh step is hardening: Place the silicon wafer on a hot plate at 200°C and heat it for 2 hours. During the hardening process, pay special attention to controlling the heating time. If the heating time is too short, it will easily lead to insufficient adhesion of the photoresist and cause the photoresist to fall off or break on the surface of the silicon wafer. If the heating time is too long, the edge of the photoresist will soften, resulting in a decrease in the resolution of the micro-groove, thereby preparing the silicon wafer master film.

[0056] For example, a silicon wafer master mold is placed in a fume hood, and trimethylchlorosilane is dripped around the silicon wafer. The hood's ventilation function is activated to remove volatile trimethylchlorosilane gas to prevent toxic damage. After the trimethylchlorosilane solidifies, the fume hood is sealed and the mold is left to stand for a first preset period of time to enhance adhesion between the photoresist and the silicon wafer. The first preset period can be 12 hours. Trimethylchlorosilane, as a release agent, can effectively reduce adhesion between PDMS and the photoresist, thereby extending the life of the master mold.

[0057] return Figure 3 S102: pouring the mixed solution of the elastic material chip onto the surface of the silicon wafer master mold for heating and curing, and peeling the elastic material chip off from the silicon wafer master mold after cooling.

[0058] For example, PDMS, a curing agent, and a surfactant (TritonX-100) are weighed in a ratio of 1000:100:1 to prepare a mixed solution of materials for an elastic material chip. Among them, the curing agent can effectively change the hardness and softness of the elastic material chip, and the surfactant is added to enhance the hydrophilicity of the surface of the elastic material chip to prevent tiny bubbles from adhering to the inside of the channel. Furthermore, before stirring the mixed solution of materials through an ultra-high-speed centrifuge, it is necessary to use a circular hole-shaped weight to balance it to ensure that both ends of the centrifuge rotor are level, and then the mixed solution of materials is placed in a vacuum dish for degassing for 30 minutes to remove bubbles in the liquid, and finally the mixed solution of materials is fully stirred through an ultra-high-speed centrifuge.

[0059] Next, place the silicon wafer master mold horizontally on a weighing dish and tape it to the areas around the silicon wafer where there are no grooves. Fix the silicon wafer master mold and the weighing dish together to make the thickness of the elastic material chip more uniform. Then slowly pour the material mixture solution from the center of the silicon wafer master mold. After the liquid level is level (that is, the material mixture solution overflows the silicon wafer master film to fill the grooves), place the weighing dish in an 80°C oven and heat for 60 minutes to cure. After heating and curing, the PDMS has been transformed from a liquid mixture into an elastic solid. After it cools, the cured elastic material chip can be peeled off from the silicon wafer master mold and cut into small pieces along the marked lines with a knife.

[0060] S103: A groove is made on the bonding plane of the elastic material chip by using a punch.

[0061] That is to say, on a plane of the elastic material chip of solidification, make groove by punch, and do not operate another plane, so that another plane of the elastic material chip is horizontally complete.And then, the plane with groove is arranged as the bonding plane of the elastic material chip.

[0062] S104: performing a bonding operation on the bonding plane of the elastic material chip and the glass sheet to prepare the microfluidic chip.

[0063] The microfluidic chip refers to the microfluidic chip proposed in the aforementioned embodiment.

[0064] Specifically, the bonding operation is performed on the bonding plane of the elastic material chip and the glass sheet in the following manner: the glass sheet is cleaned with anhydrous ethanol and deionized water, and then the cleaned glass sheet is blown dry with compressed nitrogen; the elastic material chip and the glass sheet are surface-treated with a plasma bonding machine, and after the surface treatment, the bonding plane of the elastic material chip and the surface of the glass sheet are laminated to complete the bonding.

[0065] Exemplarily, a glass sheet is placed in a staining jar and ultrasonically cleaned with anhydrous ethanol and deionized water for 10 minutes, and then the glass sheet is blown dry with compressed nitrogen. The surface of the elastic material chip and the glass sheet is then treated with oxygen plasma by a plasma bonding machine, and the treatment is performed for 30 seconds at an operating power of 30 watts (W). After oxygen plasma treatment, the bonding plane of the elastic material chip is quickly bonded to the glass sheet on both sides, and a microfluidic chip is prepared by manual bonding. After bonding is completed, the microfluidic chip is placed in an 80°C oven for one hour to enhance the bonding effect.

[0066] Among them, after the bonding plane of the elastic material chip and the glass sheet are treated with an oxygen plasma process, a large number of hydrophilic Si-OH (silicon-hydroxyl groups) are introduced into the bonding plane of the elastic material chip, and the Si-O (silicon-oxygen) chemical bonds on the surface of the glass sheet are broken and combined with the -OH hydroxyl groups in the air to form hydrophilic Si-OH (silicon-hydroxyl groups), so that the hydroxyl groups are combined when the bonding plane of the elastic material chip and the glass sheet are attached on both sides, and under heating conditions, the Si-OH (silicon-hydroxyl group) is dehydrated to form a strong Si-O-Si (silicon-oxygen-silicon bond) to achieve bonding processing.

[0067] Based on the same application concept, the embodiments of the present application also provide a fiber optic DNA sensor corresponding to the microfluidic chip provided in the above embodiments. Since the principle of solving the problem of the fiber optic DNA sensor in the embodiments of the present application is similar to that of the microfluidic chip in the above embodiments of the present application, the implementation of the fiber optic DNA sensor can refer to the implementation of the microfluidic chip, and the repeated parts will not be repeated.

[0068] For example, see Figure 5 and Figure 6 , Figure 5 Schematic diagram of the fiber optic DNA sensor provided in the embodiment of this application Figure 1 , Figure 6 Schematic diagram of the fiber optic DNA sensor provided in the embodiment of this application Figure 2 .like Figure 5 and Figure 6As shown, the fiber-optic DNA sensor includes a microfluidic chip, wherein a circular hole is provided on a glass sheet 501 of the microfluidic chip; a piezoelectric transducer 502, wherein a vibrating plate of the piezoelectric transducer is provided corresponding to the circular hole so as to expose the vibrating plate through the circular hole; a first drainage device and a second drainage device (not shown in the figure), wherein the first drainage device is connected to the first liquid flow port, and the second drainage device is connected to the second liquid flow port, so as to control the flow of liquid into and out of the liquid channel through the first drainage device and the second drainage device; and a fiber-optic DNA sensor fixed to an area of ​​the glass sheet excluding the piezoelectric transducer; wherein the bonding plane of the elastic material chip (PDMS) of the microfluidic chip is placed on the glass sheet for bonding, so that the groove of the elastic material chip is located above the fiber-optic DNA sensor and the sensing area of ​​the fiber-optic DNA sensor is located within the liquid channel, and the first fiber-optic communication port and the second fiber-optic communication port after the fiber-optic DNA sensor is placed are respectively sealed.

[0069] Specifically, a circular hole is provided in the glass sheet. The size of the hole is sufficient to fully expose the vibrating plate of the piezoelectric transducer, so that the vibrating plate is not blocked by the glass sheet. This allows the vibrating plate to vibrate fully during the vibration process, thereby increasing the flow of liquid in the liquid channel and enhancing the ability to gather bubbles at the tip and couple the bubble structure. Furthermore, after aligning the vibrating plate of the piezoelectric transducer with the circular hole of the glass sheet, ultraviolet glue is applied to the portion of the piezoelectric transducer that contacts the glass sheet, excluding the vibrating plate. The piezoelectric transducer and the glass sheet are then irradiated with ultraviolet light to securely bond.

[0070] Specifically, the fiber-optic DNA sensor is prepared by: after cleaning the surface of the fiber-optic DNA sensor to be treated with deionized water, the fiber-optic DNA sensor is immersed in a piranha solution to remove contaminants and activate hydroxyl groups on the surface of the sensing area of ​​the fiber-optic DNA sensor, and then the fiber-optic DNA sensor is washed with deionized water; after fixing the fiber-optic DNA sensor on the glass slide, the sensing area is coated with molybdenum disulfide a preset number of times to prepare the fiber-optic DNA sensor, wherein the molybdenum disulfide coating is achieved by coating the sensing area with a molybdenum disulfide aqueous solution and then placing the glass slide on a temperature control console for heating and evaporation.

[0071] The optical fiber DNA sensor is a tapered optical fiber, which is equivalent to stretching both ends of the optical fiber so that the diameters of both ends of the optical fiber DNA sensor are larger than the diameter of the sensing area in the middle of the optical fiber DNA sensor.

[0072] Before bonding the elastic material chip to the glass slide, the fiber-optic DNA sensor to be processed must undergo surface functionalization. Surface functionalization and bioconjugation are required to immobilize the DNA probe on the surface of the fiber-optic DNA sensor. The fiber-optic surface is first cleaned with deionized water. The cleaned fiber-optic DNA sensor is then immersed in a piranha solution for 40 minutes to remove any contaminants on the fiber surface and activate the -OH (hydroxyl) groups on the sensor's sensing area. The remaining piranha solution is then rinsed with deionized water. The fiber-optic DNA sensor is then affixed to the glass slide, and approximately 100 microliters (μL) of a molybdenum disulfide (MoS) aqueous solution is added to the sensing area. The glass slide is then placed on a temperature control station and heated to dryness at 80°C four times. Each time, approximately 100 microliters (μL) of the MoS aqueous solution is applied to the sensing area and heated to dryness. This process is repeated four times to achieve four MoS coatings. The MoS enhances the adhesion strength of the sensing area, thus fabricating the fiber-optic DNA sensor.

[0073] After the fiber-optic DNA sensor is completed, the glass slide with the fiber-optic DNA sensor affixed is placed under a microscope. Using the microscope, the groove of the elastic material chip is aligned with the fiber-optic DNA sensor and placed on the glass slide, ensuring that the bonding plane of the elastic material chip contacts the glass slide. The sensing area of ​​the fiber-optic DNA sensor is accurately embedded in the liquid channel of the elastic material chip, and the fiber-optic DNA sensor is embedded in the fiber-optic communication port of the elastic material chip. The first and second fiber-optic communication ports are then sealed with ultraviolet glue. This is equivalent to adding ultraviolet glue to the first and second fiber-optic communication ports, so that the portion of the fiber-optic DNA sensor embedded in the first fiber-optic communication port is adhered to the first fiber-optic communication port, and the portion of the fiber-optic DNA sensor embedded in the second fiber-optic communication port is adhered to the second fiber-optic communication port, preventing liquid in the liquid channel from entering or exiting through the first and second fiber-optic communication ports. Thus, if the length of the fiber-optic DNA sensor is greater than the length of the groove, resulting in the ends of the fiber-optic DNA sensor being outside the first and second fiber-optic communication ports, the ultraviolet glue can also seal the first and second fiber-optic communication ports.

[0074] After the fiber-optic DNA sensor is embedded in the first and second optical ports of the elastic material chip and sealed, the piezoelectric transducer's oscillating plate is aligned with the circular hole on the glass sheet and secured using UV adhesive. Furthermore, after the grooved bonding surface is bonded to the glass sheet, the non-bonding surface of the elastic material chip serves as the outer surface of the microfluidic chip. On this non-bonding surface, the first drainage device is connected to the first liquid flow port, and the second drainage device is connected to the second liquid flow port.

[0075] The drainage device includes a micro-injection needle and a hose. That is, by inserting the micro-injection needle of the first drainage device into the first liquid circulation port on a non-bonded surface, the liquid injected into the hose of the first drainage device can flow into the first liquid circulation port through the micro-injection needle and enter the liquid channel; by inserting the micro-injection needle of the second drainage device into the second liquid circulation port on a non-bonded surface, the liquid in the liquid channel can be extracted into the hose of the second drainage device through the micro-injection needle of the second drainage device, thereby achieving liquid flow into and out of the liquid channel.

[0076] Based on the same application concept, the embodiments of the present application also provide a DNA detection method corresponding to the optical fiber DNA sensor provided in the above embodiments. Since the principle of solving the problem by the DNA detection method in the embodiments of the present application is similar to that of the optical fiber DNA sensor in the above embodiments of the present application, the implementation of the DNA detection method can refer to the implementation of the optical fiber DNA sensor, and the repeated parts will not be repeated.

[0077] Specifically, the DNA detection method provided in the embodiment of the present application includes: preparing a DNA probe complementary to the DNA to be tested on the sensing area of ​​the optical fiber DNA sensor in the optical fiber DNA sensor; injecting a test solution containing the DNA to be tested into the liquid channel through the first drainage device and the second drainage device, so that the DNA to be tested and the DNA probe achieve DNA complementarity in the sensing area; collecting a spectral curve of the optical fiber DNA sensor after DNA complementation is achieved, the spectral curve being used to reflect the intensity of light corresponding to different wavelengths; and determining a target DNA concentration corresponding to the target wavelength according to the target wavelength corresponding to the characteristic peak on the spectral curve, so as to use the target DNA concentration as the DNA concentration to be tested in the test solution.

[0078] That is, a DNA probe complementary to the DNA to be tested is first prepared on the sensing area of ​​the fiber optic DNA sensor in the fiber optic DNA sensor, and then a test solution containing the DNA to be tested is injected into the liquid channel to achieve complementarity between the DNA to be tested and the DNA probe. After complementation, the spectral curve of the fiber optic DNA sensor is affected. The concentration of the DNA to be tested in the test solution is determined by analyzing the target wavelength corresponding to the characteristic peak of the spectral curve, thereby realizing the analysis of the concentration of the DNA to be tested in the test solution.

[0079] The DNA probe is prepared by: injecting a polylysine solution into the liquid channel through the first drainage device and the second drainage device, and after the polylysine solution infiltrates the optical fiber DNA sensor for a second preset time period, the optical fiber DNA sensor is cleaned with deionized water; injecting a probe solution containing a DNA probe into the liquid channel through the first drainage device and the second drainage device, and controlling the probe solution to infiltrate the optical fiber DNA sensor for a third preset time period to allow the DNA probe to adhere to the optical fiber DNA sensor, and then cleaning the optical fiber DNA sensor with TE buffer to prepare the DNA probe.

[0080] For example, see Figure 7 , Figure 7 This is a schematic diagram of the concentration analysis principle of the optical fiber DNA sensor provided in the embodiment of the present application. Figure 7 As shown, after the cleaned fiber DNA sensor is placed in the piranha solution and immersed for 40 minutes to activate the -OH (hydroxyl group) on the sensing area of ​​the fiber DNA sensor, in the process of preparing the DNA probe, the polylysine (PLL) solution is introduced into the liquid channel through one of the first drainage device and the second drainage device to fill the liquid channel with the polylysine (PLL) solution, and the fiber DNA sensor is ensured to be immersed in the PLL solution for 1 hour. Then, deionized water is injected into the liquid channel through one of the first drainage device and the second drainage device, and the polylysine (PLL) solution is simultaneously extracted from the liquid channel through the other of the first drainage device and the second drainage device to clean the fiber DNA with deionized water. The sensor is then infiltrated with a PLL solution, converting the -OH (hydroxyl) groups on the sensing region into -NH2 (amino) groups. A probe solution containing a DNA probe complementary to the DNA to be tested is then injected into the liquid channel through one of the first and second drainage devices. The probe solution is then allowed to infiltrate the fiber-optic DNA sensor for approximately one hour, allowing the -NH2 (amino) groups on the sensing region to connect with the DNA probe via NH-NH bonds. TE buffer (made from Tris and ethylenediaminetetraacetic acid (EDTA)) is then injected into the liquid channel through one of the first and second drainage devices to remove any DNA probe not bound to the -NH2 (amino) groups, leaving the DNA probe attached to the sensing region. This completes the surface modification of the ultrasonic enrichment fiber-optic DNA sensor device. Subsequent experiments require only the introduction of a test solution containing the DNA to be tested.

[0081] Then, a test solution containing the DNA to be tested is injected into the liquid channel through the first drainage device and the second drainage device, and then the piezoelectric transducer is activated to control the vibration of the vibrating plate to achieve DNA complementation between the DNA to be tested in the test solution and the DNA probes attached to the sensing area. As a result, due to the different concentrations of the DNA to be tested in different test solutions, the number of DNA probes that achieve DNA complementation on the sensing area varies, and the different numbers of DNA probes that achieve DNA complementation on the sensing area will produce different transmission spectra. Therefore, the target DNA concentration of the DNA to be tested in the test solution is analyzed by analyzing the target wavelength corresponding to the characteristic peak of the transmission spectrum.

[0082] For example, to verify the measurement stability of the fiber-optic DNA sensor, a piranha solution was injected into the liquid channel of the fiber-optic DNA sensor after DNA complementation through the first and second drainage devices. The solution was allowed to soak for 40 minutes to remove the surface structure within the sensing area. The sensing area was then functionalized using a PLL solution and probe solution of the same concentration. The sensor was then tested with PBS buffer to verify the stability of the sensor after etching with the piranha solution. Spectral measurements were repeated every hour with fresh PBS buffer, for a total of three tests. Figure 8 , Figure 8 Schematic diagram of the spectral curve of repeated tests provided in the embodiment of the present application. Figure 8 As shown, the horizontal axis wl (nm) of the spectral curve represents the wavelength in nanometers (nm), while the vertical axis level (dB) represents the power or intensity in decibels (dB); decibel is a unit that describes the ratio of power or intensity and is used to express the relative size of two quantities. In spectral measurement, decibel is usually used to represent the power or intensity of light. For example, level (dB) can represent the ratio of the measured light power to the reference power, and this ratio is expressed in decibels. The characteristic peak of the spectral curve of each test refers to the trough, and the characteristic wavelengths of each trough are 1585.84 nm, 1585.8 nm, and 1585.86 nm, with a calculated standard deviation of 0.03055, which shows that the prepared fiber optic DNA sensor has good repeatability.

[0083] Specifically, the target DNA concentration corresponding to the target wavelength is determined in the following manner: in a preliminary testing stage, spectral curves corresponding to different preset DNA concentrations are obtained to determine the characteristic wavelengths of the characteristic peaks corresponding to different preset DNA concentrations; data fitting is performed according to the characteristic wavelengths corresponding to each preset DNA concentration to determine a fitting function for reflecting the relationship between DNA concentration and wavelength; and the target DNA concentration corresponding to the target wavelength is determined using the fitting function.

[0084] Specifically, during the pre-test phase, test solutions with various known preset DNA concentrations are prepared, and DNA probes are prepared using the same polylysine (PLL) solution and probe solution, ensuring that the DNA probes in the fiber-optic DNA sensor are nearly identical. Consequently, test solutions with different preset DNA concentrations are injected into the liquid channel to obtain transmission spectra corresponding to the test solutions with the different preset DNA concentrations. Furthermore, after each test solution with a preset DNA concentration is injected into the liquid channel to obtain a transmission spectrum, a piranha solution is injected into the liquid channel via the first and second drainage devices to soak the fiber-optic DNA sensor for 40 minutes. This removes the test DNA and DNA probes that are complementary to each other on the sensor area of ​​the fiber-optic DNA sensor, thereby preventing any impact on the accuracy of the transmission spectrum corresponding to the next preset DNA concentration.

[0085] The horizontal coordinate of the spectrum curve is wavelength, the vertical coordinate is decibel, the characteristic peak refers to the data point corresponding to the minimum decibel value on the spectrum curve, and the horizontal coordinate of the characteristic peak is used as the characteristic wavelength of the spectrum curve.

[0086] See also Figure 9 , Figure 9 Schematic diagram of the spectral curves corresponding to different DNA concentrations provided in the examples of this application. Figure 9 As shown, the horizontal axis wl (nm) of the spectrum curve represents the wavelength, the unit is nanometer (nm), and the vertical axis level (dB) represents the power or intensity, the unit is decibel (dB). Figure 9 The graph shows the spectral curves obtained by the fiber optic DNA sensor in test solutions of different concentrations observed experimentally. As the DNA concentration of the test solution increases, the spectral curve of the optical fiber continues to red-shift. Specifically, the spectral curve of the DNA concentration of 1fM (femtomoles per liter) is almost indistinguishable from the spectral curve of the PBS buffer solution (equivalent to a DNA concentration of 0fM). This is because the DNA concentration of the buffer solution is too low to allow the fiber optic DNA sensor to identify the DNA to be tested in the solution. When the concentration of the DNA molecules to be tested is greater than 10fM, the right shift of the characteristic peak gradually becomes obvious, that is, the red shift of the spectrum begins to gradually become obvious. This also shows that the minimum detectable concentration of the fiber optic DNA sensor is 10fM.

[0087] See also Figure 10 , Figure 10 This is a schematic diagram of the fitting function for the relationship between DNA concentration and wavelength provided in the embodiment of the present application. In the spectral curves corresponding to different DNA concentrations, the characteristic wavelength of the characteristic peak corresponding to each DNA concentration is selected, and data fitting is performed according to each DNA concentration and its corresponding characteristic wavelength to obtain data fitting. Figure 10As shown, the abscissa of the fitting function reflects the DNA concentration, and the ordinate of the fitting function reflects the characteristic wavelength of the characteristic peak at each DNA concentration. Furthermore, when the target wavelength of the characteristic peak of the test solution is obtained through the experiment, the target DNA concentration corresponding to the target wavelength is determined using the fitting function, thereby determining the target DNA concentration of the test solution.

[0088] Moreover, the determination coefficient R of the linear fitting of the fiber optic DNA sensor under ultrasound driving with the DNA concentration increasing tenfold from 10 fM to 1 uM is 2 The value is 0.98063, indicating that the fiber-optic DNA sensor's results under ultrasound drive show a linear change within the detection concentration range. However, at concentrations above 10 nM, the relative shift trend slows significantly. This is because ultrasound enrichment prematurely saturates the capture capacity of the probe on the fiber-optic sensor surface. In other words, above 10 nM concentration, there is no excess complementary DNA on the fiber to bind to the test DNA, making it difficult to accurately detect specific concentrations even at higher test DNA concentrations.

[0089] See also Figure 11 , Figure 11 Schematic diagram of characteristic wavelengths corresponding to the presence or absence of ultrasonic drive provided in the embodiment of the present application. Figure 11 As shown in the figure, the horizontal axis is used to indicate different DNA concentrations, and the vertical axis is used to indicate the drift amount of different DNA concentrations without or with ultrasonic actuation. Different DNA concentrations include: 100fM (femtomoles per liter), 1pM (picomoles per liter), 10pM, 100pM, 1nM nanomoles per liter, 10nM, 100nM, and 1μM micromolar concentrations. Furthermore, under different DNA concentrations, ultrasonic actuation and no ultrasonic actuation are performed to determine the characteristic peak of the optical fiber DNA sensor's spectral curve. In other words, the piezoelectric transducer needs to be activated when ultrasonic actuation is performed, while the piezoelectric transducer does not need to be activated when no ultrasonic actuation is performed. In this way, for each DNA concentration, the characteristic wavelength of the characteristic peak is collected by determining whether ultrasonic actuation is performed, and the difference between the characteristic wavelength and the characteristic wavelength corresponding to the test solution with a DNA concentration of 0 is calculated. The difference is used as the drift amount, and the drift amount under no or with ultrasonic actuation is obtained for each DNA concentration.

[0090] Furthermore, if Figure 11As shown, as the DNA concentration increases, the drift of the spectral curves detected with and without ultrasonic actuation increases. Specifically, higher DNA concentrations in the test solution result in greater shifts in the characteristic peaks of the spectral curve. Furthermore, the drift with ultrasonic actuation is greater than that without ultrasonic actuation, indicating that more DNA is bound to the DNA probe under ultrasonic actuation. Consequently, the responsiveness of ultrasonic actuation is significantly higher than that without ultrasonic actuation. This demonstrates that ultrasonic actuation enhances the binding efficiency of the DNA to the fiber-optic DNA sensor, thereby increasing the sensor's sensitivity to changes in DNA concentration.

[0091] Compared with the traditional MoS2 fiber DNA sensor, this application integrates a microfluidic chip with a tip-coupled bubble structure and a piezoelectric transducer that provides ultrasonic excitation to obtain an ultrasonically driven tip-coupled bubble device, which has the advantages of strong anti-disturbance, high stability, and wide biological adaptability. Further combined with the MoS2 fiber DNA sensor, the sensitivity is improved by an order of magnitude, the detection accuracy is improved from a minimum of 100fM to a minimum of 10fM, and the responsiveness is also significantly improved. The ultrasonic enrichment enhancement structure is combined with the MoS2 fiber DNA sensor to improve the binding efficiency of the DNA to be tested in the liquid sample and the DNA probe on the sensor surface. In response to the detection needs of DNA, a MoS2 fiber DNA sensor was successfully prepared by coating a DNA probe of a specific sequence as a functional material on the surface of the optical fiber to identify its complementary chain; combining soft lithography and microfluidics technology, an ultrasonic enrichment device was made to encapsulate it, which improved the reliability and sensitivity of the sensor, and ultimately achieved low-concentration detection of the DNA molecules to be tested. The minimum detectable concentration obtained in the experiment was 10fM.

[0092] Those skilled in the art can clearly understand that, for the convenience and simplicity of description, the specific working process of the system and device described above can refer to the corresponding process in the aforementioned method embodiment, and will not be repeated here. In the several embodiments provided in this application, it should be understood that the disclosed system, device and method can be implemented in other ways. The device embodiments described above are merely schematic. For example, the division of the units is only a logical function division. There may be other division methods in actual implementation. For example, multiple units or components can be combined or integrated into another system, or some features can be ignored or not executed. Another point is that the mutual coupling or direct coupling or communication connection shown or discussed can be through some communication interfaces, indirect coupling or communication connection of devices or units, which can be electrical, mechanical or other forms.

[0093] The units described as separate components may or may not be physically separate, and the components shown as units may or may not be physical units, that is, they may be located in one place or distributed across multiple network units. Some or all of these units may be selected to achieve the purpose of this embodiment according to actual needs.

[0094] In addition, each functional unit in each embodiment of the present application may be integrated into one processing unit, or each unit may exist physically separately, or two or more units may be integrated into one unit.

[0095] If the functions are implemented in the form of software functional units and sold or used as independent products, they can be stored in a non-volatile computer-readable storage medium that is executable by a processor. Based on this understanding, the technical solution of the present application, or the part that contributes to the prior art, or the part of the technical solution, can be embodied in the form of a software product. The computer software product is stored in a storage medium and includes several instructions for enabling a computer device (which can be a personal computer, a server, or a network device, etc.) to execute all or part of the steps of the method described in each embodiment of the present application. The aforementioned storage medium includes various media that can store program codes, such as a USB flash drive, a mobile hard disk, a read-only memory (ROM), a random access memory (RAM), a magnetic disk, or an optical disk.

[0096] The above are only specific embodiments of the present application, but the scope of protection of this application is not limited thereto. Any changes or substitutions that can be easily conceived by a person skilled in the art within the technical scope disclosed in this application should be included in the scope of protection of this application. Therefore, the scope of protection of this application should be based on the scope of protection of the claims.

Claims

1. A microfluidic chip with a tip-coupled bubble structure, characterized in that: The microfluidic chip comprises a bonded elastic material chip and a glass sheet, The plane of the glass sheet covers the planar projection portion of the elastic material chip, and a plane of the elastic material chip serves as a bonding plane for bonding with the glass sheet. The bonding plane is provided with a groove running through the length direction of the elastic material chip. The groove is used to place the optical fiber biosensor in the length direction and to be filled with liquid acting on the optical fiber biosensor. The groove includes a liquid channel, a first extension area, a second extension area, and a plurality of tip-coupled bubble structures. The first extension region and the second extension region are respectively located on both sides of the liquid channel in the length direction, the multiple tip-coupled bubble structures are located on one side of the liquid channel in the width direction, the width of the liquid channel in the width direction is greater than the width of the extension region in the width direction, and the depth of the liquid channel in the depth direction is less than the depth of the extension region in the depth direction. The depth of the tip-coupled bubble structure in the depth direction is equal to the depth of the liquid channel in the depth direction. The tip-coupled bubble structure is a fan-shaped structure, one side of the fan-shaped structure is connected to the liquid channel, and the other side of the fan-shaped structure does not contact the liquid channel.

2. The microfluidic chip according to claim 1, characterized in that The first extension region includes a first optical fiber communication port and a first liquid circulation port, and the first extension region includes a second optical fiber communication port and a second liquid circulation port, wherein the first optical fiber communication port and the second optical fiber communication port are located on both sides of the liquid channel in the length direction, the first liquid circulation port is located on one side of the first optical fiber communication port in the width direction, and the second liquid circulation port is located on one side of the second optical fiber communication port in the width direction, and the specifications of the optical fiber communication ports are set according to the cross-section of the optical fiber biosensor, so that the first optical fiber communication port, the liquid channel, and the second optical fiber communication port are used to place the optical fiber biosensor in the length direction; And / or, the material of the elastic material chip includes polydimethylsiloxane, a curing agent and a surfactant.

3. A method for preparing a microfluidic chip, characterized in that: The preparation method comprises: Placing a silicon wafer master mold in a fume hood and dripping trimethylchlorosilane onto the silicon wafer master mold for a first preset period of time to seal the mold, wherein at least one groove for preparing the elastic material chip is provided on the surface of the silicon wafer master mold; Pour the mixed solution of the elastic material chip onto the surface of the silicon wafer master mold to heat and solidify it, and peel the elastic material chip from the silicon wafer master mold after cooling; Making grooves on the bonding plane of the elastic material chip by a punch; performing a bonding operation on the bonding plane of the elastic material chip and a glass sheet to prepare the microfluidic chip; Wherein, the microfluidic chip refers to the microfluidic chip as described in claim 1 or 2.

4. The preparation method according to claim 3, characterized in that The silicon wafer master mold is prepared by the following method: After cleaning the silicon wafer, place it on a heating plate and bake it; Fix the baked silicon wafer on the operating table of the spin coater, and coat the photoresist on the surface of the silicon wafer; After the coated silicon wafer is placed on a heating plate for baking, a mask is placed on the photoresist on the surface of the silicon wafer for exposure, wherein the mask includes a channel pattern corresponding to at least one of the elastic material chips; After placing the exposed silicon wafer on a heating plate for baking, the silicon wafer is developed and fixed to form at least one channel for preparing the elastic material chip on the surface of the silicon wafer; The silicon wafer is hardened to prepare the silicon wafer master mold.

5. The preparation method according to claim 3, characterized in that The bonding operation of the bonding plane of the elastic material chip and the glass sheet is performed in the following manner: The glass sheet is cleaned with anhydrous ethanol and deionized water, and then the cleaned glass sheet is blown dry with compressed nitrogen; The elastic material chip and the glass sheet are surface-treated by a plasma bonding machine, and after the surface treatment, the bonding plane of the elastic material chip and the surface of the glass sheet are attached to complete the bonding.

6. An optical fiber DNA sensor prepared in combination with molybdenum disulfide material, characterized in that: The sensor comprises: The microfluidic chip according to claim 1 or 2, wherein a circular hole is provided on the glass sheet of the microfluidic chip; a piezoelectric transducer, wherein a vibrating plate of the piezoelectric transducer is arranged corresponding to the circular hole so as to expose the vibrating plate through the circular hole; a first drainage device and a second drainage device, wherein the first drainage device is connected to the first liquid circulation port, and the second drainage device is connected to the second liquid circulation port, so as to control the flow of liquid into and out of the liquid channel through the first drainage device and the second drainage device; an optical fiber DNA sensor fixed on the glass slide in an area other than the piezoelectric transducer; Among them, the bonding plane of the elastic material chip of the microfluidic chip is placed on the glass sheet for bonding, so that the groove of the elastic material chip is located above the optical fiber DNA sensor, and the sensing area of ​​the optical fiber DNA sensor is located in the liquid channel, and the first optical fiber connecting port and the second optical fiber connecting port after the optical fiber DNA sensor is placed are sealed respectively.

7. The optical fiber DNA sensor according to claim 6, characterized in that The fiber optic DNA sensor was prepared by the following method: After cleaning the surface of the fiber optic DNA sensor to be treated with deionized water, the fiber optic DNA sensor is immersed in a piranha solution to remove contaminants and activate hydroxyl groups on the surface of the sensing area of ​​the fiber optic DNA sensor, and then the fiber optic DNA sensor is washed with deionized water; After the optical fiber DNA sensor is fixed on the glass slide, the sensing area is coated with molybdenum disulfide a preset number of times to prepare the optical fiber DNA sensor, wherein the molybdenum disulfide coating is achieved by coating the sensing area with an aqueous solution of molybdenum disulfide and then placing the glass slide on a temperature control console for heating and evaporation.

8. A DNA detection method, characterized in that: The method is applied to the optical fiber DNA sensor according to claim 6 or 7, The method comprises: preparing a DNA probe complementary to the DNA to be detected on the sensing area of ​​the optical fiber DNA sensor; Injecting a test solution containing the test DNA into the liquid channel through the first drainage device and the second drainage device, so that the test DNA and the DNA probe achieve DNA complementarity in the sensing area; Collecting a spectrum curve of the optical fiber DNA sensor after DNA complementation is achieved, wherein the spectrum curve is used to reflect the intensity of light corresponding to different wavelengths; According to the target wavelength corresponding to the characteristic peak on the spectral curve, the target DNA concentration corresponding to the target wavelength is determined, so as to use the target DNA concentration as the DNA concentration to be measured in the solution to be measured.

9. The method according to claim 8, characterized in that The DNA probe was prepared by: injecting a poly-lysine solution into the liquid channel through the first drainage device and the second drainage device, and washing the optical fiber DNA sensor with deionized water after the poly-lysine solution has soaked the optical fiber DNA sensor for a second preset period of time; A probe solution containing a DNA probe is injected into the liquid channel through the first drainage device and the second drainage device, and the probe solution is controlled to infiltrate the optical fiber DNA sensor for a third preset time period to allow the attached DNA probe to adhere to the optical fiber DNA sensor. The optical fiber DNA sensor is then cleaned with TE buffer to prepare the DNA probe.

10. The method according to claim 8, characterized in that The target DNA concentration corresponding to the target wavelength is determined by: In the pre-test stage, spectral curves corresponding to different preset DNA concentrations are obtained to determine the characteristic wavelengths of the characteristic peaks corresponding to the different preset DNA concentrations; Performing data fitting according to the characteristic wavelengths corresponding to the respective preset DNA concentrations to determine a fitting function for reflecting the relationship between DNA concentration and wavelength; The target DNA concentration corresponding to the target wavelength is determined by the fitting function.

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