A gold film-space engineering array integrated microfluidic chip and a preparation method and application thereof

Abstract

The application discloses a gold film-space engineering array integrated microfluidic chip and a preparation method and application thereof, and belongs to the technical field of microfluidic detection, and comprises a microfluidic channel, wherein the microfluidic channel comprises a sample inlet area, a detection area and a sample outlet area, the detection area is provided with a plurality of parallel detection chips; the detection chip is a gold film-space engineering arrayed SERS substrate which is modified with probe molecules; the gold film-space engineering arrayed SERS substrate comprises a polystyrene compound eye array structure layer, a nano-gold film layer and a nano-gold ball layer which are arranged from bottom to top; the polystyrene compound eye array structure layer is a monolayer film which is composed of polystyrene microspheres, and a plurality of non-overlapping pits are distributed on each polystyrene microsphere. The gold film-space engineering array integrated microfluidic chip can significantly improve the efficiency and sensitivity of analysis and detection.

Description

Technical Field

[0001] This invention relates to the field of microfluidic detection technology, specifically to a gold film-space engineering arrayed integrated microfluidic chip, its fabrication method, and its application. Background Technology

[0002] Microfluidics is a cutting-edge technology involving fluid manipulation at the microscale. It focuses on the precise control of minute amounts of fluid within channels or cavities at the micrometer or even nanometer scale. Microfluidic systems typically consist of microchannel networks, micropumps, microvalves, and other components. These tiny structures are integrated onto a chip, resembling a miniature "laboratory." Through its sophisticated channel design, microfluidics can precisely control the flow rate, direction, and mixing degree of fluids, enabling rapid sample processing. It has wide applications in numerous fields such as biomedicine, chemical analysis, and environmental science, significantly reducing reagent consumption, improving the efficiency and sensitivity of analytical detection, and opening new pathways for interdisciplinary research and practical applications.

[0003] Introducing a SERS substrate into microfluidic chips is suitable for detecting trace samples and can significantly improve detection sensitivity, enabling the accurate capture and identification of even trace-level target molecules. It also greatly enhances detection specificity, effectively distinguishing target molecules from other potential interfering substances, further improving experimental controllability and repeatability. This technology shows promising applications in multiple fields such as bioanalysis, food safety, environmental monitoring, and drug development, and is expected to drive the development of efficient, accurate, and intelligent analytical detection technologies.

[0004] Chinese patent document CN117324055A discloses a SERS microfluidic chip and its fabrication method. The SERS microfluidic chip includes a SERS substrate and a microfluidic chip. The SERS substrate is connected to the output channel end of the microfluidic chip, and the input end of the microfluidic channel is used to input the sample solution to be tested. The SERS substrate is made of porous silicon carbide material and is prepared by electrochemical deposition of silver nanoparticles and automatic adsorption of silver nanoparticles.

[0005] Chinese patent document CN118304946A discloses a microfluidic chip, its fabrication method, and a microfluidic-SERS platform. The fabrication method of the microfluidic chip includes the following steps: S1: preparing a gold nanorod array reinforced glass substrate; S2: preparing a PDMS chip including fishbone structure channels; S3: legally bonding the PDMS chip to the gold nanorod array reinforced glass substrate using oxygen plasma bonding; S4: injecting SH-PEG-biotin and streptavidin protein solutions into the flow channels of the PDMS chip, flowing through the surface of the gold nanorod array reinforced glass substrate, and performing functionalization modification.

[0006] However, the above methods have problems such as poor SERS signal stability, laser angle limitations, and inability to detect multiple targets simultaneously. Therefore, it is necessary to develop a gold film-space engineering array-integrated microfluidic chip. Summary of the Invention

[0007] This invention provides a gold film-space engineered array-integrated microfluidic chip. This microfluidic chip combines a sophisticated channel design with a gold film-space engineered array-integrated SERS substrate, enabling rapid sample detection and significantly improving the efficiency and sensitivity of analysis and detection.

[0008] The specific technical solution adopted is as follows:

[0009] A gold film-space-engineered arrayed integrated microfluidic chip includes a microfluidic channel, which comprises an inlet area, a detection area, and an outlet area. The inlet area is used to input the solution to be tested; the detection area is provided with multiple detection chips connected in parallel; and the outlet area is used to output the solution after testing.

[0010] The detection chip is a gold film-space-engineered arrayed SERS substrate modified with probe molecules. The gold film-space-engineered arrayed SERS substrate includes a polystyrene compound eye array structure layer, a gold nanofilm layer, and a gold nanosphere layer arranged from bottom to top. The thickness of the polystyrene compound eye array structure layer is 1-5 μm, the thickness of the gold nanofilm layer is 50-200 nm, and the thickness of the gold nanosphere layer is 20-50 nm. The polystyrene compound eye array structure layer is a monolayer film composed of polystyrene microspheres, and each polystyrene microsphere has several non-overlapping pits distributed on it.

[0011] Preferably, the polystyrene microspheres have a particle size of 1-5 μm, a pit depth of 50-450 nm, and a pit diameter of 100-500 nm.

[0012] Optionally, the probe molecule is at least one of DTBPA (dithiobisphenylazide), 4-ATP (4-aminothiophenol), or 4-MBA (4-mercaptobenzoic acid).

[0013] Further preferably, the detection area of ​​the microfluidic channel is provided with three parallel detection chips, namely, a gold film-space-engineered arrayed SERS substrate modified with DTBPA, a gold film-space-engineered arrayed SERS substrate modified with 4-ATP, and a gold film-space-engineered arrayed SERS substrate modified with 4-MBA.

[0014] This invention also provides a method for fabricating the aforementioned gold film-space engineering arrayed integrated microfluidic chip, comprising:

[0015] S01 designed microfluidic channels and fabricated the corresponding mold (positive mold);

[0016] In step S02, the PDMS solution is added to the mold in step S01. After curing and shaping, the detection chip is bonded to it and further encapsulated using PDMS to obtain the gold film-space engineering arrayed integrated microfluidic chip.

[0017] Optionally, the detection chip is fabricated by adding a gold film-space-engineered arrayed SERS substrate into a probe molecule solution and allowing it to stand. The concentration of the probe molecule solution is 10. -3 -10 -5 M, let stand for 30-60 minutes.

[0018] Specifically, the gold film-space-engineered arrayed SERS substrate was prepared by the following method:

[0019] S11 uses an interface self-assembly method to form a polystyrene monolayer substrate from polystyrene microspheres.

[0020] S12 loads silica nanospheres onto a polystyrene monolayer substrate, and after heating and etching to remove the silica nanospheres, a polystyrene compound eye array structure layer is obtained.

[0021] S13 prepared a gold nanofilm layer on a polystyrene compound eye array structure layer by physical sputtering, and then transferred a gold nanosphere monolayer film formed by the self-assembly of gold nanospheres to the gold nanofilm layer to obtain the gold film-space engineering arrayed SERS substrate.

[0022] Preferably, in step S11, polystyrene microspheres are self-assembled at the gas-liquid interface using an ethanol suspension, and sodium dodecyl sulfate is added as an auxiliary agent to prepare a polystyrene monolayer substrate.

[0023] Preferably, in step S12, a silica suspension is coated onto a polystyrene monolayer substrate, thereby loading silica nanospheres onto the polystyrene monolayer substrate. After heating at 100-150°C for 5-10 minutes, the silica nanospheres are removed by etching with hydrofluoric acid to form pits, thus obtaining a polystyrene compound eye array structure layer.

[0024] More preferably, the particle size of the silica nanospheres is 100-500 nm.

[0025] Further preferred, the hydrofluoric acid etching conditions are 2-8 wt% hydrofluoric acid etching for 20-60 min.

[0026] Preferably, in step S13, a gold target is used for sputtering, and the sputtering time is 5-60 min; the particle size of the gold nanospheres is 20-50 nm, and more preferably 30 nm.

[0027] The present invention also provides the application of the aforementioned gold film-space-engineered arrayed integrated microfluidic chip in the detection of hydrogen sulfide, benzaldehyde and / or putrescine.

[0028] The present invention also provides a method for detecting hydrogen sulfide, benzaldehyde and / or putrescine, comprising: using the gold film-space engineering array integrated microfluidic chip, causing the detection chip to adsorb the target molecules in the test solution, and detecting the Raman signal of the target molecules adsorbed by the detection chip by a Raman spectrometer.

[0029] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0030] (1) The present invention uses a specific gold film-space engineered arrayed SERS substrate, which includes a polystyrene compound eye array structure layer, a nano gold film layer and a nano gold ball layer arranged from bottom to top. It has high detection sensitivity, good stability and high anti-reflection ability and omnidirectional light capture, and strong SERS detection signal. Furthermore, it is integrated into a microfluidic channel, which not only has low manufacturing cost and simple detection operation process, but also enables rapid detection of samples, reduces reagent consumption, and significantly improves the efficiency and sensitivity of analysis and detection.

[0031] (2) The present invention integrates multiple SERS detection chips that have been pre-modified with different probe molecules into the same microfluidic chip in a multi-channel parallel manner, which can realize the simultaneous and rapid detection of multiple molecules in a mixed solution. Attached Figure Description

[0032] Figure 1 A schematic flowchart for fabricating a gold film-space engineering array-integrated microfluidic chip.

[0033] Figure 2 A schematic diagram of the mold used to fabricate a gold film-space engineering array-integrated microfluidic chip.

[0034] Figure 3 This is a SEM image of the polystyrene compound eye array structure layer in Example 1.

[0035] Figure 4 This is a SEM image of the gold film-space engineering arrayed SERS substrate in Example 1.

[0036] Figure 5 The image shows the SERS spectra of hydrogen sulfide, benzaldehyde, and putrescine detected by the gold film-space engineering arrayed integrated microfluidic chip in Example 1. Detailed Implementation

[0037] To make the objectives, features, and advantages of this invention more apparent and understandable, a detailed description is provided below through specific embodiments. Many specific details are set forth in the following description to provide a thorough understanding of the invention. However, the invention can be practiced in many other ways different from those described herein, and those skilled in the art can make similar modifications without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below. Technical features in various embodiments of the invention can be combined appropriately without mutual conflict.

[0038] Unless otherwise specified, the operating methods in the following examples are generally performed under conventional conditions or as recommended by the manufacturer. Contents not described in detail in this specification are prior art known to those skilled in the art. Unless otherwise specified, the experimental materials used in the examples below can be purchased from conventional biochemical reagent companies.

[0039] A schematic flowchart of the fabrication of a gold film-based arrayed integrated microfluidic chip for space engineering is shown below. Figure 1 As shown, the method includes:

[0040] (1) Polystyrene microspheres are formed into a polystyrene monolayer substrate by interface self-assembly method;

[0041] (2) Load silica nanospheres onto a polystyrene monolayer substrate, and remove the silica nanospheres by heating and etching to obtain a polystyrene compound eye array structure layer.

[0042] (3) A gold nanofilm was prepared on a polystyrene compound eye array structure layer by physical sputtering, and then a gold nanosphere monolayer formed by the self-assembly of gold nanospheres was transferred onto the gold nanofilm to obtain a gold film-space engineering arrayed SERS substrate.

[0043] (4) Modify probe molecules on a gold film-space engineered arrayed SERS substrate to prepare a detection chip;

[0044] (5) Design microfluidic channels and fabricate corresponding molds. A schematic diagram of the molds used in the following embodiments is shown below. Figure 2 As shown;

[0045] (6) Add the PDMS solution into the mold, solidify and remove it, then bond the detection chip to it, and further encapsulate it with PDMS to obtain the gold film-space engineering array integrated microfluidic chip.

[0046] Example 1

[0047] First, polystyrene (PS) microspheres (3 μm in diameter) were dispersed in ethanol to obtain a uniform ethanol suspension of polystyrene microspheres with a mass concentration of 1.25 wt%. Using a syringe, the ethanol suspension of polystyrene microspheres was injected onto an inclined glass slide (45°) at an injection rate of 10 μL / min. The suspension then flowed to the air-water interface for self-assembly. After 30 min, 200 μL of sodium dodecyl sulfate (SDS) (2.5 wt%) was added as a surfactant to further reduce the surface tension of water and change the surface properties of the PS microspheres, which is beneficial for the formation of a more stable monolayer. Subsequently, colored diffraction fringes could be observed at the air / water interface, thus preparing a polystyrene monolayer substrate. The polystyrene monolayer substrate was then transferred to an underwater silicon wafer (3 mm × 3 mm, which was subjected to 100 mW oxygen plasma treatment for 2 min before use) and then dried at room temperature.

[0048] Silica nanospheres (200 nm in diameter) were dispersed in ethanol to obtain a silica suspension (2 wt%). 2 μL of this silica suspension was added dropwise to the surface of a polystyrene monolayer substrate. The substrate was then loaded with silica nanospheres using a blade coating method with an 8° blade angle and a 30 μm gap between the blade edge and the substrate. After coating, the substrate was dried and then heated at 120 °C for 10 min. After cooling to room temperature, the substrate was immersed in a 4 wt% hydrofluoric acid solution for 30 min. After removal, the substrate was rinsed with deionized water to remove surface impurities and finally dried at room temperature to obtain a polystyrene compound eye array structure (e.g., ...). Figure 3 As shown in the figure, the polystyrene compound eye array structure layer is a monolayer film composed of polystyrene microspheres. Each polystyrene microsphere has several non-overlapping pits. The particle size of the polystyrene microspheres is 3μm, the depth of the pits is 50-200nm, and the diameter of the pit opening is 100-200nm.

[0049] A gold nanofilm was sputtered onto the surface of a polystyrene compound eye array structure for 15 minutes, resulting in uniform coverage of the pits by the gold nanofilm, which was approximately 200 nm thick. Simultaneously, 30 nm gold nanospheres self-assembled at the interface of an ethanol / n-hexane mixture and water to prepare a gold nanosphere monolayer. This monolayer was then transferred onto the gold nanofilm to prepare the gold film-space-engineered arrayed SERS substrate (e.g., [example needed]). Figure 4 (As shown).

[0050] The gold film-space-engineered arrayed SERS substrate obtained in this embodiment includes a polystyrene compound eye array structure layer, a gold nanofilm layer, and a gold nanosphere layer arranged from bottom to top. The thickness of the polystyrene compound eye array structure layer is about 3 μm, the thickness of the gold nanofilm layer is about 200 nm, and the thickness of the gold nanosphere layer is about 30 nm.

[0051] Gold film-space engineering arrayed SERS substrates were added to a concentration of 10... -4 After standing in DTBPA, 4-ATP or 4-MBA solution for 40 min, gold film-space engineered arrayed SERS substrate modified with DTBPA, gold film-space engineered arrayed SERS substrate modified with 4-ATP, and gold film-space engineered arrayed SERS substrate modified with 4-MBA were obtained.

[0052] The microfluidic channel was designed and the corresponding mold was fabricated. Specifically, the mold can fabricate a gold film-space engineering array integrated microfluidic chip with an overall length of 52.8830 mm, a width of 22.2000 mm, and a height of 3.0000 mm. The microfluidic channel is 0.5-1 mm wide and includes a sample inlet area, a detection area, and a sample outlet area. The sample inlet area is used to input the solution to be detected, and the sample outlet area is used to output the solution after detection. The detection area has a square structure with a side length of 2.5-3.5 mm and a height of 0.8-1.2 mm.

[0053] A PDMS mixture with a mass ratio of 10:1 (PDMS main agent to curing agent) was added to the above mold. After curing and molding at 80°C, the mold was removed. Then, gold film-space engineered arrayed SERS substrates modified with DTBPA, 4-ATP, and 4-MBA were bonded in parallel to the detection area. After further encapsulation with PDMS, the gold film-space engineered arrayed integrated microfluidic chip was obtained.

[0054] Example 2

[0055] The steps and parameters in this embodiment are the same as those in Embodiment 1. The only difference is that silica suspensions are prepared using silica nanospheres with particle sizes of 100, 300, 400, and 500 nm, respectively, and finally, a gold film-space engineering array-integrated microfluidic chip is obtained.

[0056] Example 3

[0057] The steps and parameters in this embodiment are the same as those in Embodiment 1. The only difference is that gold nanosphere monolayer films are assembled using gold nanospheres with particle sizes of 10, 20, 40, and 50 nm, respectively, and then a gold film-space engineering array-integrated microfluidic chip is further prepared.

[0058] Sample Analysis

[0059] Figure 5The SERS spectra of hydrogen sulfide, benzaldehyde, and putrescine were detected using the gold film-space engineering arrayed integrated microfluidic chip described in Example 1. As can be seen from the figure, using hydrogen sulfide solution as the test solution, the measured SERS signal is at 1137 cm⁻¹. -1 The peak value at 1620 cm⁻¹ was significantly enhanced, indicating that hydrogen sulfide was successfully detected; using benzaldehyde solution as the test solution, the measured SERS spectrum was at 1620 cm⁻¹. -1 A new characteristic peak appeared at 715 cm⁻¹, indicating that benzaldehyde was successfully detected; using putrescine solution as the test solution, the measured SERS spectrum was at 715 cm⁻¹. -1 A new characteristic peak appears at 690 cm⁻¹. -1 The decrease in the peak value indicates that benzaldehyde was successfully detected.

[0060] The embodiments described above provide a detailed explanation of the technical solutions of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Any modifications, additions, or similar substitutions made within the scope of the principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A gold film-space engineering arrayed integrated microfluidic chip, characterized in that, It includes a microfluidic channel, which comprises an injection zone, a detection zone, and an output zone. The injection zone is used to input the solution to be tested; the detection zone is equipped with multiple parallel detection chips; and the output zone is used to output the solution after testing. The detection chip is a gold film-space-engineered arrayed SERS substrate modified with probe molecules. The gold film-space-engineered arrayed SERS substrate includes a polystyrene compound eye array structure layer, a gold nanofilm layer, and a gold nanosphere layer arranged from bottom to top. The thickness of the polystyrene compound eye array structure layer is 1-5 μm, the thickness of the gold nanofilm layer is 50-200 nm, and the thickness of the gold nanosphere layer is 20-50 nm. The polystyrene compound eye array structure layer is a monolayer film composed of polystyrene microspheres, and each polystyrene microsphere has several non-overlapping pits distributed on it.

2. The gold film-space engineering arrayed integrated microfluidic chip according to claim 1, characterized in that, The polystyrene microspheres have a particle size of 1-5 μm, a pit depth of 50-450 nm, and a pit diameter of 100-500 nm.

3. The gold film-space engineering arrayed integrated microfluidic chip according to claim 1, characterized in that, The probe molecule is at least one of DTBPA, 4-ATP, or 4-MBA.

4. The method for fabricating a gold film-space engineering arrayed integrated microfluidic chip according to any one of claims 1-3, characterized in that, include: S01 designed microfluidic channels and fabricated the corresponding molds; In step S02, the PDMS solution is added to the mold in step S01. After curing and shaping, the detection chip is bonded to it and further encapsulated using PDMS to obtain the gold film-space engineering arrayed integrated microfluidic chip.

5. The method for fabricating a gold film-space engineering arrayed integrated microfluidic chip according to claim 4, characterized in that, The fabrication method of the detection chip is as follows: a gold film-space-engineered arrayed SERS substrate is added to a probe molecule solution and allowed to stand. The concentration of the probe molecule solution is 10. -3 -10 -5 M, let stand for 30-60 minutes.

6. The method for fabricating a gold film-space engineering arrayed integrated microfluidic chip according to claim 5, characterized in that, The gold film-space-engineered arrayed SERS substrate was prepared by the following method: S11 uses an interface self-assembly method to form a polystyrene monolayer substrate from polystyrene microspheres. S12 loads silica nanospheres onto a polystyrene monolayer substrate, and after heating and etching to remove the silica nanospheres, a polystyrene compound eye array structure layer is obtained. S13 prepared a gold nanofilm layer on a polystyrene compound eye array structure layer by physical sputtering, and then transferred a gold nanosphere monolayer film formed by the self-assembly of gold nanospheres to the gold nanofilm layer to obtain the gold film-space engineering arrayed SERS substrate.

7. The method for fabricating a gold film-space engineering arrayed integrated microfluidic chip according to claim 6, characterized in that, In step S11, polystyrene microspheres are self-assembled at the gas-liquid interface using an ethanol suspension, and sodium dodecyl sulfate is added as an auxiliary agent to prepare a polystyrene monolayer substrate.

8. The method for fabricating a gold film-space engineering arrayed integrated microfluidic chip according to claim 6, characterized in that, In step S12, a silica suspension is coated onto a polystyrene monolayer substrate, thereby loading silica nanospheres onto the polystyrene monolayer substrate. After heating at 100-150°C for 5-10 minutes, the silica nanospheres are removed by etching with hydrofluoric acid to form pits, resulting in a polystyrene compound eye array structure layer.

9. The application of the gold film-space-engineered arrayed integrated microfluidic chip according to any one of claims 1-3 in the detection of hydrogen sulfide, benzaldehyde and / or putrescine.

10. A method for detecting hydrogen sulfide, benzaldehyde, and / or putrescine, characterized in that, include: Using the gold film-space engineering arrayed integrated microfluidic chip as described in any one of claims 1-3, the detection chip adsorbs the target molecules in the test solution, and the Raman signal of the target molecules adsorbed by the detection chip is detected by a Raman spectrometer.

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