Electrochemical micro-fluidic chip for simultaneously detecting multiple bacteria
By integrating a three-electrode system on a microfluidic chip, multiple bacteria can be detected simultaneously using electrochemical signals at different potentials. This solves the problem of the inability to detect multiple bacteria simultaneously in existing technologies, and improves detection efficiency and specificity.
Patent Information
- Application Number
- CN202423003552.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Utility models(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-05
- Publication Date
- 2025-11-11
- Estimated Expiration
- 2034-12-05
AI Technical Summary
Existing electrochemical microfluidic chips cannot simultaneously detect multiple bacteria in a sample passing through the same separation chamber, resulting in low detection efficiency.
A three-electrode system is integrated on the substrate of a microfluidic chip. Multiple target bacteria in the sample are separated by a separation chamber, and electrochemical signals are obtained at different potentials using the three-electrode system, enabling the simultaneous detection of multiple bacteria.
It enables the simultaneous detection of multiple bacteria, improving the sensitivity and specificity of detection and solving the problem that existing technologies cannot detect multiple bacteria simultaneously.
Smart Images
Figure CN223538816U_ABST
Abstract
Description
Technical Field
[0001] This utility model relates to the field of microfluidic chip technology, specifically to an electrochemical microfluidic chip that can simultaneously detect multiple bacteria. Background Technology
[0002] With food safety issues becoming increasingly serious, rapid and accurate detection of foodborne pathogens has become an important issue of global concern.
[0003] In the prior art, by combining electrochemical biosensors with microfluidic chips, efficient bacterial capture and detection can be achieved on a small chip. However, existing electrochemical microfluidic chips cannot detect multiple bacteria in a sample passing through the same separation chamber at the same time, resulting in low detection efficiency.
[0004] Therefore, how to achieve simultaneous detection of multiple bacteria based on the same separation chamber remains a technical challenge that urgently needs to be solved. Utility Model Content
[0005] (a) Technical problems to be solved
[0006] To address the shortcomings of existing technologies, this invention provides an electrochemical microfluidic chip for the simultaneous detection of multiple bacteria, solving the problem that existing electrochemical microfluidic chips cannot simultaneously detect multiple bacteria in samples passing through the same separation chamber.
[0007] (II) Technical Solution
[0008] To achieve the above objectives, this utility model provides the following technical solution:
[0009] In this invention, an electrochemical microfluidic chip for simultaneous detection of multiple bacteria is used for sample detection, comprising a substrate and microfluidic channels for sample flow and processing.
[0010] The microfluidic channel integrates an electrochemical electrode, which is a three-electrode system comprising a working electrode, a reference electrode, and a counter electrode, for detecting electrochemical signals.
[0011] The microfluidic channel includes a separation chamber, a mixing channel, and an electrochemical detection chamber integrated with an electrochemical electrode. The separation chamber and the mixing channel are connected and flow with the electrochemical detection chamber.
[0012] The end of the mixing channel away from the separation chamber is connected to a connecting pipe three. The end of the connecting pipe three away from the mixing channel is provided with multiple inlets, and each inlet is connected to the connecting pipe three through a corresponding connecting pipe.
[0013] After separation in the separation chamber, the sample flows through the three-electrode system for detection via the first connecting tube, and then flows out through the outlet after being connected to the second connecting tube.
[0014] The microfluidic chip also includes a micropump for driving sample flow;
[0015] During detection, characteristic peak values of the corresponding target bacteria in the sample at different potentials are obtained.
[0016] Furthermore, the mixing channel is serpentine or spiral-shaped.
[0017] Furthermore, the separation chamber is connected to the connecting pipe 1 via a three-way pipe, the first port of the three-way pipe is in communication with the separation chamber, the second port of the three-way pipe is in communication with the connecting pipe 1, and a solution injection port is provided on the third port of the three-way pipe.
[0018] The inlet is used to inject the buffer solution into the separation chamber through the three-way tube.
[0019] Furthermore, the width of the microfluidic channel ranges from 10 to 500 micrometers, and the height of the microfluidic channel ranges from 10 to 200 micrometers.
[0020] Furthermore, the material of the microfluidic channel is polydimethylsiloxane.
[0021] (III) Beneficial Effects
[0022] This invention provides an electrochemical microfluidic chip for the simultaneous detection of multiple bacteria. Compared with existing technologies, it has the following advantages:
[0023] By integrating a three-electrode system onto the substrate of a microfluidic chip, multiple target bacteria in a sample can simultaneously flow through a separation chamber for separation. Then, by utilizing the electrochemical signals obtained by the three-electrode system at different potentials, multiple target bacteria can be detected simultaneously, solving the problem in existing technologies that cannot simultaneously detect multiple bacteria in a sample passing through the same separation chamber. Attached Figure Description
[0024] To more clearly illustrate the technical solutions in the embodiments of this utility model or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this utility model. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.
[0025] Figure 1 This is a schematic diagram of the structure of an electrochemical microfluidic chip for the simultaneous detection of multiple bacteria.
[0026] Figure 2This is a schematic diagram illustrating the process of using an electrochemical microfluidic chip to simultaneously detect multiple bacteria.
[0027] Figure label:
[0028] 1. Substrate; 11. Separation chamber; 12. T-junction; 121. Inlet; 122. Connecting tube one; 123. Connecting tube two; 124. Outlet; 13. Mixing channel; 131. Connecting tube three; 132. Inlet; 2. Three-electrode system. Detailed Implementation
[0029] To make the objectives, technical solutions, and advantages of the embodiments of this utility model clearer, the technical solutions in the embodiments of this utility model are described clearly and completely. Obviously, the described embodiments are only some embodiments of this utility model, not all embodiments. Based on the embodiments of this utility model, all other embodiments obtained by those skilled in the art without creative effort are within the protection scope of this utility model.
[0030] This application provides an electrochemical microfluidic chip for the simultaneous detection of multiple bacteria, which solves the problem that existing electrochemical microfluidic chips cannot simultaneously detect multiple bacteria in a sample passing through the same separation chamber, and at the same time improves the sensitivity of the detection process.
[0031] The technical solution in this application is to solve the above-mentioned technical problems, and the general idea is as follows:
[0032] In existing technologies, microfluidic chip technology utilizes fluid control in micron or nanometer-scale channels to achieve precise sample processing and analysis. This technology has advantages such as small sample requirements, fast reaction speed, and easy integration, and is widely used in fields such as biomedical detection and environmental monitoring. However, single microfluidic chip technology cannot simultaneously detect multiple bacteria in a sample passing through the same separation chamber, resulting in certain limitations in the sensitivity and specificity of bacterial detection, especially when simultaneously detecting multiple bacteria, it is difficult to achieve ideal results.
[0033] Research has found that, for example Figures 1-2 As shown, by integrating a three-electrode system onto the substrate of a microfluidic chip, multiple target bacteria in a sample can simultaneously flow through a separation chamber for separation. Then, by utilizing the electrochemical signals obtained by the three-electrode system at different potentials, multiple target bacteria can be detected simultaneously, solving the problem in existing technologies that it is impossible to detect multiple bacteria in a sample passing through the same separation chamber at the same time.
[0034] To better understand the above technical solutions, the following will provide a detailed explanation of the technical solutions in conjunction with the accompanying drawings and specific implementation methods.
[0035] like Figures 1-2 As shown, an electrochemical microfluidic chip for simultaneous detection of multiple bacteria is used for sample detection, including a substrate 1 and microfluidic channels for sample flow and processing.
[0036] The microfluidic channel integrates an electrochemical electrode, which is a three-electrode system 2. The three-electrode system 2 includes a working electrode, a reference electrode, and a counter electrode, and is used to detect electrochemical signals.
[0037] The microfluidic channel includes a separation chamber 11, a mixing channel 13, and an electrochemical detection chamber integrated with an electrochemical electrode. The separation chamber 11 and the mixing channel 13 are connected and flow with the electrochemical detection chamber.
[0038] The mixing channel 13 is connected to a connecting pipe 131 at the end away from the separation chamber 11. The connecting pipe 131 is provided with multiple inlets 132 at the end away from the mixing channel 13. Each inlet 132 is connected to the connecting pipe 131 through a corresponding connecting pipe.
[0039] After being separated in the separation chamber 11, the sample flows through the three-electrode system 2 via the connecting tube 122 for detection, and after being connected to the connecting tube 2 123, it flows out through the outlet 124.
[0040] The microfluidic chip also includes a micropump for driving sample flow;
[0041] During detection, characteristic peak values of the corresponding target bacteria in the sample at different potentials are obtained.
[0042] It should be noted that the electrochemical signals at different potentials originate from the different redox potential characteristic peaks of different metal ions or electrochemically active molecules loaded in the sodium alginate gel microsphere probes that specifically bind to specific target bacteria, thereby enabling the simultaneous electrochemical detection of multiple bacteria.
[0043] By integrating the three-electrode system 2 onto the substrate 1 of the microfluidic chip, multiple target bacteria in the sample flow through the separation chamber 11 simultaneously for separation. Then, the electrochemical signals obtained by the three-electrode system 2 at different potentials are used to achieve simultaneous detection of multiple target bacteria, solving the problem in the prior art that it is impossible to detect multiple bacteria in a sample passing through the same separation chamber 11 at the same time.
[0044] like Figure 1 As shown, the mixing channel 13 is serpentine or spiral-shaped. The serpentine or spiral shape enhances the degree of liquid reaction during sample processing and strengthens the specificity of the initial sample treatment.
[0045] like Figure 1As shown, the separation chamber 11 is connected to the connecting pipe 122 via a three-way pipe 12. The first port of the three-way pipe 12 is connected to the separation chamber 11, the second port of the three-way pipe 12 is connected to the connecting pipe 122, and a solution injection port 121 is provided on the third port of the three-way pipe 12.
[0046] Inlet 121 is used to inject buffer solution into separation chamber 11 through three-way tube 12.
[0047] By setting up the three-way tube 12, it is easy to inject the buffer solution into the separation chamber 11. The buffer solution does not need to pass through the mixing channel 13 and can directly enter the separation chamber 11, which speeds up the time for the buffer solution to enter the separation chamber 11, thereby making it easier to wash away unbound bacteria, excess electrochemically active probes and other impurities.
[0048] Specifically, the buffer solution is PBS buffer.
[0049] like Figure 2 As shown, the width of the microfluidic channel ranges from 10 to 500 micrometers, and the height of the microfluidic channel ranges from 10 to 200 micrometers. This allows the fluid in the chip channel to maintain a certain flow rate and velocity, ensuring smooth sample delivery and facilitating the fabrication of the microfluidic chip.
[0050] Specifically, the material of the microfluidic channel is polydimethylsiloxane (PDMS).
[0051] Fabrication process of microfluidic chips:
[0052] Step 1: Fabricate microfluidic channels on substrate 1 of the microfluidic chip;
[0053] Step 2: Integrate electrochemical electrodes within the microfluidic channel;
[0054] Fabrication of electrochemical microfluidic chip: Microfluidic channels were fabricated on polydimethylsiloxane (PDMS) substrate 1 using photolithography, and a three-electrode system 2 was integrated in the channel, including Ag / AgCl as reference electrode, carbon as working electrode (4 mm in diameter) and carbon as auxiliary electrode.
[0055] like Figure 2 As shown, the application examples of microfluidic chips for simultaneous bacterial detection are as follows (detection of Salmonella Typhimurium and Escherichia coli):
[0056] Step 1: Inject the sample to be tested, the corresponding magnetic beads for capturing and detecting target bacteria, and the corresponding sodium alginate gel microspheres into the microfluidic channel through the corresponding inlet 132, respectively. The sample may be a food sample, a medical sample, or an environmental sample.
[0057] Step 2: Capture the corresponding target bacteria in the sample using the corresponding magnetic beads;
[0058] The target bacteria are Salmonella typhimurium and Escherichia coli.
[0059] Step 3: Bind the corresponding sodium alginate gel microspheres loaded with metal ions or electrochemically active molecules to the captured target bacteria;
[0060] Step 4: Simultaneous detection of multiple target bacteria is achieved by using electrochemical signals obtained at different potentials through electrochemical electrodes.
[0061] Wherein: the electrochemical signal is measured by cyclic voltammetry or differential pulse voltammetry; the analysis of the electrochemical signal is performed by data processing and result output using specific software.
[0062] When modifying antibodies with magnetic beads: The specific antibody is immobilized on the surface of the magnetic beads using the EDC / NHS conjugation method. First, the magnetic beads are suspended in MES buffer, and EDC and NHS are added to activate the surface of the magnetic beads. Then, the antibody solution is added and reacted at room temperature to achieve the binding of the antibody to the magnetic beads. Finally, the magnetic beads are washed with PBS buffer to remove unbound antibodies.
[0063] When modifying antibodies and electrochemically active molecules using sodium alginate gel microspheres: Sodium alginate gel microspheres are prepared using the sol-gel method, and different metal ions or electrochemically active molecules, as well as specific antibodies, are immobilized through cross-linking reactions. The specific steps include: first, mixing sodium alginate solution with different metal ions or electrochemically active molecules, then adding it dropwise to CaCl2 solution to form gel microspheres and immobilize the electrochemically active components; then, suspending the microspheres in solutions containing different antibodies to react and modify the antibodies; finally, washing the gel microsphere probes with PBS buffer.
[0064] It should be noted that the particle size of the magnetic beads ranges from 0.1 to 10 micrometers;
[0065] The particle size range of the sodium alginate gel microspheres is 10-200 micrometers;
[0066] The loaded metal ions are lead ions, copper ions, zinc ions, or cadmium ions, etc.
[0067] Other electrochemically active molecules loaded are thionine, Prussian blue, Nile blue, methylene blue, or ferrocene, etc.
[0068] Bacterial capture and detection: The sample to be tested is injected into the microfluidic channel, and the sample flow is driven by a micropump. Antibody-modified magnetic beads are introduced into the microfluidic channel and specifically bind to the target bacteria in the sample to form an antibody-bacterial complex. Subsequently, sodium alginate gel microspheres loaded with different metal ions or electrochemically active molecules and modified with specific antibodies are introduced into the channel to further bind with the antibody-bacterial complex to form a sandwich structure. Electrochemical signals are detected at different potentials by electrochemical electrodes. Based on the characteristic peak values of the electrical signals, the simultaneous detection of multiple target bacteria can be achieved.
[0069] System optimization: The modification process of magnetic beads and gel microspheres was optimized to improve antibody modification efficiency and gel microsphere stability. The potential range and scan rate of electrochemical detection were optimized to obtain the best electrochemical signal. By selecting different metal ions or electrochemically active molecules and utilizing their characteristic peaks at different potentials, simultaneous detection of multiple target bacteria was achieved. Mathematical modeling and signal processing techniques were used to analyze and decode the detected electrochemical signals, improving the accuracy of multiplex detection.
[0070] Wide range of applications: This technology can be widely used in fields such as food safety testing, medical diagnosis, and environmental monitoring.
[0071] It should be noted that two different sodium alginate gel microspheres need to be prepared separately, each modified and immobilized with different specific antibodies (Salmonella typhimurium antibody and Escherichia coli antibody) and different metal ions (lead ions or copper ions, etc.). These microspheres bind to Salmonella typhimurium and Escherichia coli, respectively. Two types of magnetic beads are also prepared, each modified with the two different specific antibodies mentioned above, for the recognition and capture of different target bacteria. The corresponding sodium alginate gel microspheres can only be modified and immobilized with the corresponding specific antibodies, and the corresponding magnetic beads can only recognize and capture the corresponding target bacteria.
[0072] In summary, compared with existing technologies, it has the following beneficial effects:
[0073] 1. By integrating the three-electrode system 2 onto the substrate 1 of the microfluidic chip, multiple target bacteria in the sample flow through the separation chamber 11 simultaneously for separation. Then, the electrochemical signals obtained by the three-electrode system 2 at different potentials are used to achieve simultaneous detection of multiple target bacteria, which solves the problem in the prior art that it is impossible to detect multiple bacteria in the sample passing through the same separation chamber 11 at the same time.
[0074] 2. By making the mixing channel 13 serpentine or spiral, the bending of the serpentine or spiral shape can improve the degree of liquid reaction during sample processing and enhance the specificity of the initial sample processing.
[0075] 3. By setting up the three-way tube 12, it is easy to inject the buffer solution into the separation chamber 11. The buffer solution does not need to pass through the mixing channel 13 and can directly enter the separation chamber 11, which speeds up the time for the buffer solution to enter the separation chamber 11, thereby making it easier to wash away unbound bacteria, excess electrochemically active probes and other impurities.
[0076] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.
[0077] The above embodiments are only used to illustrate the technical solutions of this utility model, and are not intended to limit it. Although this utility model has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of this utility model.
Claims
1. An electrochemical microfluidic chip for simultaneous detection of multiple bacteria, used for sample detection, characterized in that, Includes a substrate (1) and microfluidic channels for sample flow and processing; The microfluidic channel integrates an electrochemical electrode, which is a three-electrode system (2). The three-electrode system (2) includes a working electrode, a reference electrode, and a counter electrode, and is used to detect electrochemical signals. The microfluidic channel includes a separation chamber (11), a mixing channel (13), and an electrochemical detection chamber with integrated electrochemical electrodes. The separation chamber (11) and the mixing channel (13) are connected and flow with the electrochemical detection chamber. The mixing channel (13) is connected to a connecting pipe three (131) at one end away from the separation chamber (11). The connecting pipe three (131) is provided with multiple inlets (132) at one end away from the mixing channel (13). Each inlet (132) is connected to the connecting pipe three (131) through a corresponding connecting pipe. After separation in the separation chamber (11), the sample flows through the three-electrode system (2) via the first connecting tube (122) for detection, and after connecting to the second connecting tube (123), it flows out through the outlet (124). The microfluidic chip also includes a micropump for driving sample flow; During detection, characteristic peak values of the corresponding target bacteria in the sample at different potentials are obtained.
2. The electrochemical microfluidic chip for simultaneous detection of multiple bacteria as described in claim 1, characterized in that, The mixing channel (13) is serpentine or spiral.
3. The electrochemical microfluidic chip for simultaneous detection of multiple bacteria as described in claim 1, characterized in that, The separation chamber (11) is connected to the connecting pipe (122) through a three-way pipe (12). The first port of the three-way pipe (12) is connected to the separation chamber (11), the second port of the three-way pipe (12) is connected to the connecting pipe (122), and a solution injection port (121) is provided on the third port of the three-way pipe (12). The inlet (121) is used to inject the buffer solution into the separation chamber (11) through the three-way tube (12).
4. The electrochemical microfluidic chip for simultaneous detection of multiple bacteria as described in claim 1, characterized in that, The width of the microfluidic channel ranges from 10 to 500 micrometers, and the height of the microfluidic channel ranges from 10 to 200 micrometers.
5. The electrochemical microfluidic chip for simultaneous detection of multiple bacteria as described in claim 1, characterized in that, The material of the microfluidic channel is polydimethylsiloxane.