Microfluidic electrical stimulation chip device

Abstract

The utility model provides a microfluidic electrical stimulation chip device, which comprises a substrate, an outside electrode tube, an inside electrode, a sealing tube and an electrode screw, the substrate comprises a flow channel cavity, a liquid inlet hole, a power-on hole and a liquid outlet hole, and a flow channel and the liquid inlet hole, the power-on hole and the liquid outlet hole which are communicated to the flow channel cavity are formed in the side surface of the substrate; the outer diameter of the outer side electrode tube is consistent with the aperture of the substrate micro-channel; the outer diameter value of the inner electrode is lower than the inner diameter value of the outer electrode tube; the outer diameter of the sealing pipe is consistent with the aperture of the substrate runner, and the inner diameter of the sealing pipe is consistent with the outer diameter of the inner side electrode; and the electrode screw is embedded into the electrifying hole of the substrate. When the micro-fluidic electrical stimulation chip device provided by the utility model is used for preparing exosomes, cell suspension can form microliter-level micro-fluid between the inner electrode tube and the outer electrode tube, and cells continuously pass through the micro-fluid between the inner electrode tube and the outer electrode tube one by one and are stimulated by electrical signals between the inner electrode tube and the outer electrode tube at the same time by adjusting the size of a micro-channel, so that the cells are promoted to secrete exosomes, and the exosomes are prepared. The yield of the exosome is improved.

Description

Technical Field

[0001] The utility model relates to the fields of medicine, biology and microfluidics, and in particular to a microfluidics electrical stimulation chip device. Background Art

[0002] Exosomes are tiny vesicles secreted from cells to the outside of cells. The diameter of these tiny vesicles is generally between 30-150nm and they are coated by a phospholipid bilayer. Exosomes carry a variety of biomarkers such as proteins, mRNA and miRNA, and are directly involved in important life activities such as intercellular information transmission, antigen transmission, protein transport and RNA transcription. They are closely related to the occurrence of various diseases such as cancer, so they have great application value in the exploration of the pathogenesis of diseases and the detection of related diseases.

[0003] The production of exosomes mainly depends on the natural secretion during the culture of living cells, and the yield is low. Studies have shown that changes in cell culture conditions, including changes in serum, pH, temperature, oxygen content, culture additives, or providing electrical stimulation, can promote the release of exosomes from cells. For example, US2022243191A1 discloses a method for producing exosomes, which includes applying radio frequency (RF) electrical stimulation with a frequency of 0.05 to 5 MHz to cells and culturing cells to enhance the production and secretion of exosomes in cells. However, the overall electrical stimulation intensity of this cell culture chamber is weak, the electric field distribution is uneven, and the effect of promoting cell secretion of exosomes is general. The amount of exosomes secreted by HEK293 cells is only 2.3 times.

[0004] Microfluidics is an emerging science and technology for manipulating fluids in microscale space. As a carrier for realizing microfluidics technology, microfluidic chips have the characteristics of continuity, miniaturization, integration, and low consumption. They can integrate the basic operation units involved in biochemical reactions, such as sample preparation, reaction, separation, and detection, into a chip of a few square centimeters or even smaller. However, there are currently no reports on the use of microfluidics to promote cell secretion of exosomes.

[0005] The present invention provides a microfluidic electrical stimulation chip device that combines electrical stimulation with microfluidics. Through this device, precise and online microfluidic electrical stimulation of cells at the micron level can be achieved, which greatly improves the exosome yield and production efficiency and can be better suitable for industrial production. Utility Model Content

[0006] The utility model provides a microfluidic electrical stimulation chip device for promoting cells to secrete exosomes.

[0007] The utility model provides a microfluidic electrical stimulation chip device, comprising a substrate 1, an outer electrode tube 2, an inner electrode 3, a sealing screw 4, and an electrode screw 5, and is characterized in that: the substrate 1 comprises a flow channel cavity 11, a liquid inlet hole 12, a power-through hole 13 and a liquid outlet hole 14, wherein: the flow channel cavity 11 and the liquid inlet hole 12, the power-through hole 13 and the liquid outlet hole 14 connected to the flow channel cavity 11 are opened on the side of the substrate 1; the outer electrode tube 2 is a hollow cylinder, and its outer diameter is consistent with the aperture of the flow channel cavity 11; the inner electrode 3 is a cylinder, and its outer diameter is lower than the inner diameter of the outer electrode tube 2; the outer diameter of the sealing screw 4 is consistent with the aperture of the flow channel cavity 11, and the inner diameter of the sealing screw 4 is consistent with the outer diameter of the inner electrode 3; the electrode screw 5 is a conductive screw embedded in the power-through hole 13 of the substrate 1 and energized to the outer electrode tube 2.

[0008] in:

[0009] The length of the outer electrode tube 2 is 2 to 40 mm;

[0010] The inner electrode 3 has a length of 10 to 80 mm, and its outer diameter is 0.2 to 5.0 mm lower than the inner diameter of the outer electrode tube 2. The inner electrode 3 can be a hollow or solid cylinder.

[0011] The sealing screw 4 has a length of 1 to 20 mm;

[0012] The outer electrode tube 2, the inner electrode 3 and the electrode screw 5 can be made of one of iron, copper, stainless steel, zinc, nickel, titanium, platinum, gold or graphite, preferably copper, titanium, platinum, and more preferably titanium.

[0013] The microfluidic electrical stimulation chip device provided by the utility model can be used to prepare exosomes by the following method: a cell or cell microcarrier suspension enters the microfluidic electrical stimulation chip device through the liquid inlet 12, flows through the microchannel between the inner and outer electrodes at a stable flow rate, and flows out from the liquid outlet 14. The inner and outer electrode tubes can pass direct current or alternating current.

[0014] in:

[0015] The stable flow rate is 0.05 to 50.00 mL / min, preferably 5 to 10 mL / min;

[0016] The voltage of the direct current is 10-60V, and the voltage of the alternating current is 10-60V with a frequency of 50Hz.

[0017] The voltage between the inner and outer electrode tubes forms an electric field perpendicular to the flow direction of the microfluid between the electrode tubes.

[0018] The utility model provides a microfluidic electrical stimulation chip device, which can apply different degrees of electrical stimulation to the cells passing through by adjusting the voltage when the device is used to promote the secretion of exosomes by cells.

[0019] When preparing exosomes using the microfluidic electrical stimulation chip device provided by the utility model, cells or cell microcarrier suspensions can form microliter-level microfluids between the inner and outer electrodes. By adjusting the size of the microchannel, cells can pass through continuously, one by one, and in sequence, while receiving electrical signal stimulation between the inner and outer electrodes, thereby promoting cell secretion of exosomes and increasing exosome production. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Schematic diagram of microfluidic electrical stimulation chip device:

[0021] In the figure: substrate 1, flow channel cavity 11, liquid inlet hole 12, liquid outlet hole 14, outer electrode tube 2, inner electrode 3, sealing screw 4, electrode screw 5.

[0022] Figure 2 Schematic diagram of substrate 1 in the microfluidic electrical stimulation chip device:

[0023] In the figure: flow channel cavity 11, liquid inlet hole 12, power supply hole 13, liquid outlet hole 14.

[0024] Figure 3 Particle size distribution diagram of test sample 1 in Test Example 1.

[0025] Figure 4 Particle size distribution diagram of test sample 2 in Test Example 1.

[0026] Figure 5 Particle size distribution diagram of test sample 3 in Test Example 1.

[0027] Figure 6 Particle size distribution diagram of test sample 4 in Test Example 1.

[0028] Figure 7 Particle size distribution diagram of test sample 5 in Test Example 1.

[0029] Figure 8 Particle size distribution diagram of the control sample in Experimental Example 1.

[0030] Fig. 9 Particle size distribution diagram of the blank sample in Experimental Example 1.

[0031] Fig.10 Particle size distribution diagram of the test sample in Test Example 3.

[0032] Fig.11 Particle size distribution diagram of the control sample in Experimental Example 3. DETAILED DESCRIPTION

[0033] The present invention is further described by way of the following examples, which are not intended to limit the present invention in any way.

[0034] The chip specifications of all microfluidic electrical stimulation chip devices involved in the following embodiments are expressed as (inner diameter of outer electrode tube 2 - outer diameter of inner electrode 3)*length of outer electrode tube 2, and the unit is millimeter (mm).

[0035] Example 1 Microfluidic electrical stimulation chip device

[0036] like Figure 1 and Figure 2 As shown, the microfluidic electrical stimulation chip device provided by the utility model comprises a substrate 1, a flow channel cavity 11, a liquid inlet hole 12, a liquid outlet hole 14, an outer electrode tube 2, an inner electrode 3, a sealing screw 4, and an electrode screw 5, wherein: a hole is drilled on the side of the substrate 1 to form the flow channel cavity 11; the substrate 1 is tapped to form a liquid inlet hole 12, a power-on hole 13 and a liquid outlet hole 14 and connected to the flow channel cavity 11; the outer electrode tube 2 is a hollow cylinder with a length of 2 to 40 mm, inserted into the flow channel cavity 11 of the substrate 1 and aligned with the liquid inlet hole 12 and the liquid outlet hole 14, and the power-on hole 13 is formed on the surface of the substrate 1. 3, an electrode screw 5 is screwed into the outer electrode tube 2 to contact and fix the outer electrode tube 2; both ends of the flow channel cavity 11 are sealed and fixed by sealing screws 4 with through holes, and the length of the sealing screws 4 is 1 to 20 mm; the inner electrode 3 is a hollow or solid cylinder with a length of 10 to 80 mm, and its outer diameter is 0.2 to 5.0 mm lower than the inner diameter of the outer electrode tube 2. The inner electrode 3 is inserted into the sealing screw 4 and sealed and fixed; the outer electrode tube 2, the inner electrode 3 and the electrode screw 5 can be made of one of iron, copper, stainless steel, zinc, nickel, titanium, platinum, gold or graphite.

[0037] Experimental Example 1: Promoting exosome secretion from 293F cells using a microfluidic electrical stimulation chip device

[0038] 1. Materials and Equipment:

[0039] Microfluidic electrical stimulation chip device (1):

[0040] A circular flow channel cavity 11 is drilled on the side of the substrate 1 and opened up, with a diameter of 4mm. M5-type threads are made at both ends of the flow channel cavity 11. M3-type liquid inlet holes 12, power holes 13, and liquid outlet holes 14 are tapped on the substrate 1 to open up to the flow channel cavity 11; the outer electrode tube 2 is a hollow cylinder with an outer diameter of 3mm, a wall thickness of 0.5mm, a length of 10mm, and is made of platinum. It is inserted into the flow channel cavity 11 of the substrate 1 and aligned with the liquid inlet hole 12 and the liquid outlet hole 14. A copper electrode screw 5 is screwed into the power hole 13 to contact and fix the outer electrode tube 2; both ends of the flow channel cavity 11 are sealed and fixed with M5-type sealing screws 4 with 1mm through holes; the inner electrode 3 is a cylinder with an outer diameter of 1mm and a length of 50mm. It is made of platinum. The inner electrode 3 is inserted into the sealing screw 4 and sealed and fixed to obtain specifications Microfluidic electrical stimulation chip device with platinum electrodes.

[0041] Microfluidic electrical stimulation chip device (2):

[0042] A circular flow channel cavity 11 is drilled on the side of the substrate 1 and opened up, with a diameter of 4mm. M5-type threads are made at both ends of the flow channel cavity 11. M3-type liquid inlet holes 12, power holes 13, and liquid outlet holes 14 are tapped on the substrate 1 to open up to the flow channel cavity 11; the outer electrode tube 2 is a hollow cylinder with an outer diameter of 4mm, a wall thickness of 0.5mm, a length of 10mm, and is made of titanium. It is inserted into the flow channel cavity 11 of the substrate 1 and aligned with the liquid inlet hole 12 and the liquid outlet hole 14. A copper electrode screw 5 is screwed into the power hole 13 to contact and fix the outer electrode tube 2; both ends of the flow channel cavity 11 are sealed and fixed with M5-type sealing screws 4 with 1mm through holes; the inner electrode 3 is a cylinder with an outer diameter of 2mm and a length of 50mm. It is made of titanium. The inner electrode 3 is inserted into the sealing screw 4 and then sealed and fixed to obtain specifications. Microfluidic electrical stimulation chip device with electrodes made of titanium.

[0043] Suspension 293F cells: Thermo Fisher Scientific (China), catalog number: A14527.

[0044] Suspension 293F cells for the experiment: 200 mL of suspension 293F cells in the logarithmic growth phase were taken, with a concentration of 2.0×10 6 cells / mL, and diluted to 1.5×10 cells / mL using chemically defined medium (Situofan, catalog number: SH31037). 6 cells / mL.

[0045] Tabletop refrigerated centrifuge: Thermo Fisher, model: ST1R Plus.

[0046] Sterile hydrophilic polyethersulfone filter membrane: Millipore, product number: SLGPR33RB.

[0047] Ultracentrifuge: Thermo Fisher, model: Sorvall WX100+.

[0048] PBS: Solebol, catalog number: P1020.

[0049] Nanoparticle tracking analyzer: Malvern, model: NS300.

[0050] Vertical electrophoresis tank: Bio-Rad, model: 1658001.

[0051] Electrophoresis instrument power supply: Bio-Rad, model: 1658034.

[0052] Peristaltic pump: Lange Constant Flow Pump Co., Ltd., model: L100-1S-1.

[0053] Voltage stabilizer: Zhejiang Chint Electric Co., Ltd., model: TND1-0.5.

[0054] AC voltage regulator: Shanghai Delixi Switch Co., Ltd., model: TDGC3-500W.

[0055] 2. Sample preparation:

[0056] Test sample:

[0057] Use a peristaltic pump to pump 20mL of suspended 293F cells for the experiment into the microfluidic electrical stimulation chip device at a flow rate of 5mL / min. Use an AC voltage regulator connected to a voltage regulator to set the voltage to 10V, 15V, 20V, 40V, and 60V in sequence to energize the inner and outer electrodes. After the cells are electrically stimulated in the microchannel between the inner and outer electrodes, the cells are recovered. The recovered cells are cultured for 48 hours, and the culture supernatant is collected for subsequent exosome extraction and detection. The specific test samples are shown in Table 1:

[0058] Table 1 Test sample list

[0059] Control sample:

[0060] Take 20 mL of suspended 293F cells for the experiment, culture them under the same conditions for 48 h, and collect the culture supernatant for exosome extraction and detection.

[0061] Blank sample:

[0062] Take 20 mL of suspended 293F cells for the experiment, centrifuge to remove the cells, take the supernatant and culture it under the same conditions for 48 hours, and collect the culture supernatant for exosome extraction and detection.

[0063] 3. Isolation and characterization of exosomes:

[0064] The cell culture supernatants of the test samples, control samples, and blank samples were collected respectively, and differential ultracentrifugation was performed to separate and extract exosomes, and the method was as follows: first, centrifugation was performed at a centrifugal force of 300 g for 10 minutes to remove the cell precipitate; then, centrifugation was performed at a centrifugal force of 2000 g for 10 minutes to remove the cell debris precipitate; then, centrifugation was performed at a centrifugal force of 10000 g for 20 minutes to remove the precipitates such as microvesicles and apoptotic bodies, and the supernatant was collected; the supernatant was filtered through a 0.2 μm sterile hydrophilic polyethersulfone filter membrane and the filtrate was collected; the filtrate was centrifuged at a centrifugal force of 100000 g for 70 minutes, the supernatant was discarded, the precipitate was resuspended with 35 mL PBS, and the supernatant was discarded after centrifugation again at a centrifugal force of 100000 g for 70 minutes, and the precipitate was collected with 100 μL PBS to obtain exosomes.

[0065] The obtained exosome samples were compared and characterized in terms of quantity and particle size, and the exosome production increase multiple under different voltage conditions in the microfluidic electrical stimulation chip device was calculated.

[0066] The number and size of exosomes were measured using a nanoparticle tracking analyzer: an appropriate amount of exosomes was taken and diluted with PBS, the nanoparticle tracking analyzer was turned on, 1 mL of sample was drawn with a 1 mL sterile syringe, and it was slowly injected into the injector, the camera screen was observed, the brightness and focus were adjusted, the syringe was installed in the syringe pump, and the parameters were set before the injection measurement was started. The nanoparticle tracking analyzer records the number of nanoparticles and the Brownian motion trajectory by capturing the scattered light of nanoscale particles in the sample, and after obtaining the Brownian motion rate of the nanoparticles, the particle size was calculated using the Stokes-Einstein equation to obtain the number and size of exosomes obtained from different samples.

[0067] The exosome particle size and concentration obtained from different samples are shown in Table 2. The exosome production multiple calculated according to the ratio of the number of exosomes secreted by the test sample and the control sample is shown in Table 3, where: the corrected exosome concentration is the sample exosome concentration of the test sample or the control sample minus the sample exosome concentration of the blank sample, so as to deduct the exosomes secreted by the cells before the experiment; the number of exosomes secreted by the cells is the corrected exosome concentration divided by the cell concentration; the distribution diagram of the exosome particle size of different samples is shown in Figure 3-Figure 9 shown.

[0068] Table 2 Exosome size and concentration obtained from different samples

[0069]

[0070] Table 3 Exosome production increase of different samples

[0071]

[0072] 4. Conclusion

[0073] Experimental data show that compared with control samples and blank samples, the use of a microfluidic electrical stimulation chip device can promote 293F cells to secrete more exosomes and significantly increase exosome production.

[0074] Experimental Example 2 Effect of different specifications of microfluidic electrical stimulation chip devices on the production of exosomes in 293F cells

[0075] The sources and culture of the 293F cells, experimental suspension 293F cells, peristaltic pump, voltage stabilizer, and AC voltage regulator involved in this experiment are the same as those in Experimental Example 1.

[0076] Microfluidic electrical stimulation chip device Figure 1 and Figure 2 As shown, in this experiment, microfluidic electrical stimulation chips of different specifications are obtained by changing the diameter of the flow channel cavity 11, the inner diameter of the outer electrode tube 2, the length of the outer electrode tube 2, and the outer diameter of the inner electrode 3.

[0077] Use a peristaltic pump to transfer approximately 1.5 × 10 6 The suspended 293F cells used in the cells / mL experiment were pumped into microfluidic electrical stimulation chip devices of different specifications at a flow rate of 10 mL / min. The AC voltage regulator connected to the voltage stabilizer was used to set the voltage to 30 V to energize the inner and outer electrodes. The cells were electrically stimulated in the microchannel between the inner and outer electrodes. After the cells were recovered, they were cultured for 48 hours, and the culture supernatant was collected. The exosomes were separated and extracted for counting and characterization. The separation, extraction and characterization methods were the same as those in Experimental Example 1. The calculation of the exosome production increase of 293F cells after electrical stimulation by microfluidic electrical stimulation chip devices of different specifications is shown in Table 4.

[0078] Table 4 Exosome production increase of 293F cells obtained by microfluidic electrical stimulation chips of different specifications

[0079]

[0080] in conclusion:

[0081] The utility model provides a method for preparing exosomes using a microfluidic electrical stimulation chip device. Under the conditions of 30V and 10mL / min, microfluidic electrical stimulation chip devices of different specifications can stimulate 293F cells to secrete more exosomes.

[0082] Experimental Example 3: Promoting the secretion of exosomes by mesenchymal stem cells using a microfluidic electrical stimulation chip device

[0083] 1. Materials and Equipment:

[0084] Microfluidic electrical stimulation chip device: a circular flow channel cavity 11 is drilled on the side of the substrate 1 and opened up to a diameter of 4mm. M5-type threads are made at both ends of the flow channel cavity 11. M3-type liquid inlet holes 12, power holes 13, and liquid outlet holes 14 are tapped on the substrate 1 to open up to the flow channel cavity 11; the outer electrode tube 2 is a hollow cylinder with an outer diameter of 4mm, a wall thickness of 1mm, and a length of 10mm. It is made of titanium and is inserted into the flow channel cavity 11 of the substrate 1 to align with the liquid inlet hole 12 and the liquid outlet hole 14. A copper electrode screw 5 is screwed into the power hole 13 to contact and fix the outer electrode tube 2; both ends of the flow channel cavity 11 are sealed and fixed with M5-type sealing screws 4 with 1mm through holes; the inner electrode 3 is a cylinder with an outer diameter of 1mm and a length of 50mm. It is made of platinum and is sealed and fixed after the inner electrode 3 is inserted into the sealing screw 4. Get the flow channel specifications Microfluidic Dionex stimulation chip device with titanium electrodes.

[0085] Benchtop refrigerated centrifuge: Thermo Fisher, ST1R Plus.

[0086] Sterile hydrophilic polyethersulfone membrane: Millipore, SLGPR33RB.

[0087] Ultra-high speed centrifuge: Thermo Fisher, Sorvall WX100+.

[0088] PBS: Solebol, P1020.

[0089] Nanoparticle tracking analyzer: Malvern, NS300.

[0090] Vertical electrophoresis tank: Bio-Rad, 1658001.

[0091] Electrophoresis instrument power supply: Bio-Rad, 1658034.

[0092] Umbilical cord mesenchymal stem cells (MSC): sourced from Shandong Qilu Stem Cell Engineering Co., Ltd.

[0093] MSC cells used in the experiment:

[0094] MSCs were seeded onto microcarriers and placed in a bioreactor (Beijing Huakan Biotechnology Co., Ltd., model: 3D The cells were cultured in miniSPIN at a stirring speed of 33 rpm until the third day, the culture supernatant was removed, and fresh culture medium (Beijing Huakan Biotechnology Co., Ltd., catalog number: RMQ010) was replaced.

[0095] 2. Sample preparation:

[0096] Test sample:

[0097] 100 mL of experimental MSC cell microcarrier suspension was stirred at 50 rpm in a stirred bioreactor, and the microcarrier suspension was pumped into the microfluidic electrical stimulation chip B at a flow rate of 10 mL / min using a peristaltic pump. The AC voltage regulator connected to the voltage regulator was adjusted to 20 V to energize the inner and outer electrodes, and the cells were electrically stimulated in the microchannel between the inner and outer electrodes. After the cells were recovered, they were cultured in a carbon dioxide shaking incubator at a stirring speed of 33 rpm for 48 hours, and the cell culture supernatant was collected for subsequent exosome extraction and detection.

[0098] Control sample:

[0099] 100 ml of experimental MSC cell microcarrier suspension was added to a stirred bioreactor and cultured under the same conditions for 48 h. The cell culture supernatant was collected for subsequent exosome extraction and detection.

[0100] 3. Isolation and Characterization of Exosomes

[0101] The cell culture supernatants of the test samples and the control samples were collected respectively, and the exosomes were separated and extracted by differential ultracentrifugation. The specific method was as follows: centrifugation at a centrifugal force of 300g for 10 minutes to remove the cell precipitate; centrifugation at a centrifugal force of 2000g for 10 minutes to remove the cell debris precipitate; centrifugation at a centrifugal force of 10000g for 20 minutes to remove the precipitate such as microvesicles and apoptotic bodies, and the supernatant was collected; the supernatant was filtered through a 0.2μm sterile hydrophilic polyethersulfone filter membrane and the filtrate was collected; the filtrate was centrifuged at a centrifugal force of 100000g for 70 minutes, the supernatant was discarded, the precipitate was resuspended with 35mL PBS, and the supernatant was discarded after centrifugation at a centrifugal force of 100000g for 70 minutes, and the precipitate was collected with 100μL PBS to obtain exosomes.

[0102] The quantity and particle size of the obtained exosome samples were compared and characterized, and the exosome production increase multiple under microfluidic electrical stimulation conditions was calculated.

[0103] The number and size of exosomes were determined using a nanoparticle tracking analyzer. Take an appropriate amount of exosomes and dilute them with PBS, turn on the nanotracking analyzer, draw 1 mL of sample with a 1 mL sterile syringe, slowly push it into the injector, observe the camera screen, adjust the brightness and focus, install the syringe into the syringe pump, set the parameters and start the injection measurement. The nanoparticle tracking analyzer records the number of nanoparticles and the Brownian motion trajectory by capturing the scattered light of nanoscale particles in the sample, obtains the Brownian motion rate of the nanoparticles, and calculates the particle size through the Stokes-Einstein equation, and finally obtains the number and size of exosomes.

[0104] The particle size and concentration of exosomes obtained from different samples are shown in Table 5. The exosome production multiple calculated according to the ratio of the number of exosomes secreted by the test sample and the control sample is shown in Table 6. The particle size distribution of exosomes from different samples is shown in Figure 10-11 shown.

[0105] Table 5 Exosome particle size and concentration obtained from different samples

[0106]

[0107] Table 6 Exosome production increase folds of different samples

[0108]

[0109] 4. Conclusion

[0110] It can be seen from the experimental data that compared with the control sample, the microfluidic electrical stimulation chip device provided by the utility model can greatly promote the secretion of exosomes by MSC cells, and can increase the amount of exosomes secreted by a single cell to 122.5 times.

Claims

1. A microfluidic electrical stimulation chip device, comprising a substrate (1), an outer electrode tube (2), an inner electrode (3), a sealing screw (4), and an electrode screw (5), wherein: The substrate (1) comprises a flow channel cavity (11), a liquid inlet hole (12), a power-on hole (13) and a liquid outlet hole (14), wherein: the flow channel cavity (11) and the liquid inlet hole (12), the power-on hole (13) and the liquid outlet hole (14) connected to the flow channel cavity (11) are opened on the side of the substrate (1); the outer electrode tube (2) is a hollow cylinder, and its outer diameter is consistent with the hole diameter of the flow channel cavity (11); the inner electrode (3) is a cylinder, and its outer diameter is lower than the inner diameter of the outer electrode tube (2); the outer diameter of the sealing screw (4) is consistent with the hole diameter of the flow channel cavity (11), and the inner diameter of the sealing screw (4) is consistent with the outer diameter of the inner electrode (3); and the electrode screw (5) is a conductive screw embedded in the power-on hole (13) of the substrate (1) and energized to the outer electrode tube (2).

2. The microfluidic electrical stimulation chip device according to claim 1, characterized in that: The length of the outer electrode tube (2) is 2 to 40 mm.

3. The microfluidic electrical stimulation chip device according to claim 1, characterized in that: The length of the inner electrode (3) is 10 to 80 mm, and its outer diameter is 0.2 to 5 mm lower than the inner diameter of the outer electrode tube (2).

4. The microfluidic electrical stimulation chip device according to claim 3, characterized in that: The inner electrode (3) is a hollow or solid cylinder.

5. The microfluidic electrical stimulation chip device according to claim 1, characterized in that: The length of the sealing screw (4) is 1 to 20 mm.

6. The microfluidic electrical stimulation chip device according to claim 1, characterized in that: The outer electrode tube (2), the inner electrode (3) and the electrode screw (5) are made of one of iron, copper, stainless steel, zinc, nickel, titanium, platinum, gold or graphite.

7. The microfluidic electrical stimulation chip device according to claim 1, characterized in that: The outer electrode tube (2), the inner electrode (3) and the electrode screw (5) are made of one of copper, titanium or platinum.

8. The microfluidic electrical stimulation chip device according to claim 7, characterized in that: The outer electrode tube (2), the inner electrode (3) and the electrode screw (5) are made of titanium.

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