Microfluidic Impedance Detection Chip for Particle Counting

Abstract

The present invention discloses a microfluidic impedance detection chip for particle counting, comprising an upper substrate and a lower substrate bonded to each other, the lower substrate being provided with an inlet reservoir, a curved inertial focusing channel, and a particle detection assembly, the two ends of the curved inertial focusing channel being connected to the inlet reservoir and the particle detection assembly, respectively; the curved inertial focusing channel being enclosed by an outer boundary and an inner boundary, the outer boundary being in the shape of a circular arc, the inner boundary being in the shape of an Archimedean spiral, so that the width of the curved inertial focusing channel decreases along the direction from the inlet reservoir to the particle detection assembly; the upper substrate being provided with a liquid inlet and a liquid outlet, the liquid inlet being connected to the inlet reservoir, and the liquid outlet being connected to the particle detection assembly. The chip of the present invention is small in size, easy to carry, has a high detection throughput, is low in cost, and is simple to manufacture, and has great application potential in counting high-concentration tiny particles, impedance detection, and the like.

Description

Technical Field

[0001] The present invention relates to the field of microfluidic technology, and in particular to a microfluidic impedance detection chip for particle counting. Background Art

[0002] Impedance counting of micron-sized particles in liquids plays a crucial role in clinical diagnosis, genetic engineering, metallurgy, environmental protection, and other fields. Gao Lingeng's proposed impedance counting method for micron-sized particles consists of an inlet reservoir, an upper detection electrode, a lower detection electrode, and an outlet reservoir. This method, which uses the pressure threshold that the solution must break through as it passes through a rapidly narrowing microchannel, precisely positions the low-melting-point detection electrode within the chip. This method, which then performs counting, has attracted considerable attention for its low cost, small size, and high signal-to-noise ratio.

[0003] During Gao Lingeng's capillary restriction valve test electrode fabrication process, in order to precisely control the pressure threshold required for the low-melting-point liquid electrode material to break through the capillary restriction valve, expensive and complex experimental equipment such as a hot plate, air compressor, air source triplex, proportional control valve, and solenoid valve were required. Furthermore, Gao Lingeng independently designed and developed a corresponding air pressure configuration system, which then carried out melting, extraction, injection, and inspection of the electrode material. This method clearly involved a complex fabrication process and high production costs.

[0004] In the capillary restriction valve detection chip proposed by Gao Lingeng, a particle count of 15-micron diameter particles was performed using a particle throughput of 3 particles per 100 seconds to prevent clogging of the detection aperture. This structure clearly exhibits low detection throughput, making it difficult to count particles at higher concentrations. Summary of the Invention

[0005] Based on the technical problems existing in the background technology, the present invention proposes a microfluidic impedance detection chip for particle counting. The chip is small in size, easy to carry, has high detection throughput, low cost and is easy to manufacture. It has great application potential in counting tiny particles at higher concentrations, impedance detection and other aspects.

[0006] The microfluidic impedance detection chip for particle counting proposed in the present invention includes an upper substrate and a lower substrate that are bonded to each other, wherein the lower substrate is provided with an inlet liquid reservoir, a curved inertial focusing flow channel and a particle detection component, and the two ends of the curved inertial focusing flow channel are respectively connected to the inlet liquid reservoir and the particle detection component; the curved inertial focusing flow channel is surrounded by an outer boundary and an inner boundary to form a flow channel, the outer boundary is an arc shape, and the inner boundary is an Archimedean spiral shape, so that the width of the curved inertial focusing flow channel decreases along the direction from the inlet liquid reservoir to the particle detection component; the upper substrate is provided with a liquid inlet hole and a liquid outlet hole, the liquid inlet hole is connected to the inlet liquid reservoir, and the liquid outlet hole is connected to the particle detection component.

[0007] Preferably, the particle detection assembly comprises an outlet reservoir connected to an end of the curved inertial focusing channel away from the inlet reservoir, and the outlet reservoir is connected to a power supply electrode, an upper detection electrode and a lower detection electrode.

[0008] Preferably, the power electrode includes a power electrode liquid reservoir and a power electrode filling channel, the power electrode filling channel is used to connect the power electrode liquid reservoir and the outlet liquid reservoir, the upper substrate is provided with a power electrode liquid inlet hole, and the power electrode liquid inlet hole is connected to the power electrode liquid reservoir.

[0009] Preferably, the upper detection electrode includes an upper detection electrode liquid reservoir and an upper detection electrode filling channel, the upper detection electrode filling channel is used to connect the upper detection electrode liquid reservoir and the outlet liquid reservoir, the upper substrate is provided with an upper detection electrode liquid inlet hole, and the upper detection electrode liquid inlet hole is connected to the upper detection electrode liquid reservoir.

[0010] Preferably, the lower detection electrode includes a lower detection electrode liquid reservoir and a lower detection electrode filling channel, the lower detection electrode filling channel is used to connect the lower detection electrode liquid reservoir and the outlet liquid reservoir, the upper substrate is provided with a lower detection electrode liquid inlet hole, and the lower detection electrode liquid inlet hole is connected to the lower detection electrode liquid reservoir.

[0011] Preferably, a plurality of evenly distributed focusing bosses are provided on a side of the inner boundary close to the outer boundary.

[0012] Preferably, an impurity filtering micro-column array is provided in the inlet liquid storage tank.

[0013] Beneficial technical effects of the present invention:

[0014] (1) The microfluidic inertial focusing impedance detection chip proposed in the present invention can highly focus the particles when passing through the detection connection hole due to the curved inertial focusing flow channel, thereby solving the problem of clogging of the detection connection hole due to excessive detection flux of the particles when using the capillary restriction valve electrode counting method. The capillary restriction valve detection chip can perform particle counting detection at a rate of 30 particles per second, which is a significant improvement compared with the detection flux of 3 particles per 100 seconds of the original detection chip. The outer boundary of the curved inertial focusing flow channel of the present invention is an arc shape, and the inner boundary is an Archimedean spiral shape, so that the width of the curved inertial focusing flow channel changes gradually. Compared with the equal-width focusing flow channel, the secondary flow intensity of the present invention is greater and the focusing effect is better. The design of the focusing boss in the curved inertial focusing flow channel can further improve the focusing effect.

[0015] (2) The method for fabricating the capillary restriction valve detection electrode proposed in the present invention only involves basic experimental equipment such as a hot plate, a vacuum dryer, and an injection pump, which greatly reduces the production cost of the detection electrode and eliminates the steps required to design and configure the corresponding intelligent injection system, thereby simplifying the electrode production steps. The prepared chip is small in size, easy to carry, has a high detection flux, is low in cost, and is easy to fabricate. It has great application potential in counting small particles (such as biological cells) at high concentrations and impedance detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 This is a schematic diagram of the structure of the microfluidic impedance detection chip proposed in the present invention;

[0017] Figure 2 This is an exploded view of the microfluidic impedance detection chip proposed in the present invention;

[0018] Figure 3 A schematic structural diagram of the lower substrate proposed in the present invention;

[0019] Figure 4 Schematic diagram of the inlet liquid storage tank and impurity filtering micro-column array proposed by the present invention;

[0020] Figure 5 This is a schematic diagram of the structure of the curved inertial focusing flow channel proposed in the present invention;

[0021] Figure 6 This is a schematic diagram of the structure of the particle detection component proposed by the present invention;

[0022] Figure 7 This is a schematic diagram of the capillary restriction valve and detection orifice structure proposed by the present invention;

[0023] Figure 8 for Figure 4 The secondary flow vector distribution diagram generated in the cross section at the end BB of the mid-bend inertial focusing flow channel;

[0024] Figure 9 The secondary flow vector distribution diagram generated in the end cross section of the medium-width focusing flow channel in the comparative example;

[0025] Figure 10 The waveform detected in Example 1 within 0.2 seconds;

[0026] Figure 11 for Figure 10 The waveform shows the signal obtained by removing the 0.7 mV noise signal;

[0027] Figure 12 This is the processed particle signal diagram.

[0028] In the figure: 1-upper substrate, 2-lower substrate, 3-liquid inlet, 4-inlet liquid reservoir, 41-impurity filtering microcolumn array, 5-curved inertial focusing channel, 51-outer boundary, 52-inner boundary, 6-focusing boss, 7-power electrode, 71-power electrode liquid reservoir, 72-power electrode filling channel, 73-power electrode capillary restriction valve, 74-power electrode connecting hole, 8-upper detection electrode, 81-upper detection electrode liquid reservoir, 82-upper detection electrode filling channel, 83-upper detection electrode capillary restriction valve, 84-upper detection electrode connecting hole, 9-lower detection electrode, 91-lower detection electrode liquid reservoir, 92-lower detection electrode filling channel, 93-lower detection electrode capillary restriction valve, 94-lower detection electrode connecting hole, 10-outlet liquid reservoir, 11-liquid outlet, 12-power electrode liquid inlet hole, 13-upper detection electrode liquid inlet hole, 14-lower detection electrode liquid inlet hole. DETAILED DESCRIPTION

[0029] The present invention will be further explained below with reference to specific embodiments.

[0030] Example

[0031] Reference Figure 1-3 and Figure 5 The microfluidic impedance detection chip for particle counting proposed in the present invention is made of polydimethylsiloxane (PDMS) through a soft photolithography process. The process specifically includes steps such as photolithography of a SU-8 template, PDMS casting, and PDMS-PDMS bonding, and has the advantages of high processing precision. The chip includes an upper substrate 1 and a lower substrate 2 that are bonded to each other. The lower substrate 2 is provided with an inlet reservoir 4, a curved inertial focusing channel 5, and a particle detection component. The two ends of the curved inertial focusing channel 5 are respectively connected to the inlet reservoir 4 and the particle detection component. The curved inertial focusing channel 5 is surrounded by an outer boundary 51 and an inner boundary 52. ​​The outer boundary 51 is in the shape of a circular arc, and the inner boundary 52 is in the shape of an Archimedean spiral. This allows the width of the curved inertial focusing channel 5 to decrease from the inlet reservoir 4 to the particle detection component. The upper substrate 1 is provided with a liquid inlet hole 3 and a liquid outlet hole 11. The liquid inlet hole 3 is connected to the inlet reservoir 4, and the liquid outlet hole 11 is connected to the particle detection component.

[0032] Reference Figure 3 The particle detection component includes an outlet liquid reservoir 10 connected to the end of the curved inertial focusing channel 5 away from the inlet liquid reservoir 4. The outlet liquid reservoir 10 is connected to the power electrode 7, the upper detection electrode 8 and the lower detection electrode 9. The filling material in the electrode flow channel of the power electrode 7, the upper detection electrode 8 and the lower detection electrode 9 is a low-melting-point conductive bismuth indium tin alloy.

[0033] Reference Figure 6 and Figure 7The power electrode 7 includes a power electrode liquid reservoir 71 and a power electrode filling channel 72. The power electrode filling channel 72 is used to connect the power electrode liquid reservoir 71 and the outlet liquid reservoir 10. The upper substrate 1 is provided with a power electrode liquid inlet hole 12, and the power electrode liquid inlet hole 12 is connected to the power electrode liquid reservoir 71.

[0034] The upper detection electrode 8 includes an upper detection electrode liquid reservoir 81 and an upper detection electrode filling channel 82. The upper detection electrode filling channel 82 is used to connect the upper detection electrode liquid reservoir 81 and the outlet liquid reservoir 10. The upper substrate 1 is provided with an upper detection electrode liquid inlet hole 13, and the upper detection electrode liquid inlet hole 13 is connected to the upper detection electrode liquid reservoir 81.

[0035] The lower detection electrode 9 includes a lower detection electrode liquid reservoir 91 and a lower detection electrode filling channel 92. The lower detection electrode filling channel 92 is used to connect the lower detection electrode liquid reservoir 91 and the outlet liquid reservoir 10. The upper substrate 1 is provided with a lower detection electrode liquid inlet hole 14, and the lower detection electrode liquid inlet hole 14 is connected to the lower detection electrode liquid reservoir 91.

[0036] The capillary restriction valve structure (73, 83, 93) of the present invention is a structure with arc-shaped sides and a rapidly narrowed width, which provides a pressure threshold when the molten electrode material is pressed into the capillary restriction valve structure (73, 83, 93). When the pressure is less than the threshold, the molten material cannot pass through the structure, thereby achieving the purpose of accurately controlling the position of the material; the upper and lower detection holes (84, 94) are used to reduce the cross-sectional area of ​​the water flow at the hole to increase the resistance, thereby increasing the target particle signal under the same current change.

[0037] Reference Figure 3 and Figure 5 A plurality of evenly distributed focusing bosses 6 are provided on the side of the inner boundary 52 close to the outer boundary 51 .

[0038] Reference Figure 4 An impurity filtering micro-column array 41 is provided in the inlet liquid storage tank 4 for filtering large impurities.

[0039] During the electrode fabrication process, a hot plate is used to convert the solid 60° low-melting-point bismuth indium tin electrode material into a liquid state. A section of the bubble-free liquid material is extracted using a plastic hose and then allowed to stand (repeat the operation to obtain multiple sections of hose). A piston is used to block the chip liquid inlet (3). Three sections of hose containing alloy material are inserted into the inlet of the capillary restriction valve pipe (71, 81, 91). Then, one end of a hollow hose is inserted into the chip liquid outlet (11). The other end is connected to a syringe pump to form a closed microspace.

[0040] The connected chip is placed in an 80°C vacuum dryer and allowed to stand for one minute to allow the solidified alloy material to melt again. The syringe pump extraction mode is turned on and extraction is performed at a rate of 200 μL / s for 15 seconds. The air in the microspace is extracted from the liquid outlet (11) to reduce the internal pressure of the microspace. The pressure difference between atmospheric pressure and the microspace is used to press the liquid conductive material in the plastic hose to the small holes (74, 84, 94) at the terminal of the capillary restriction valve structure (73, 83, 93).

[0041] After the extraction is completed, the chip is taken out, and the inserted hose is removed after the chip cools down. The chip is then placed on a high-temperature hot plate for 1 minute. DuPont wires are inserted at the electrode pipe inlets (71, 81, 91) for connecting to external circuits during testing. The filling status of the liquid electrode is observed under a microscope. No bubbles or obvious fractures appear, and a successfully prepared detection electrode is finally obtained.

[0042] During the implementation process, a metal drainage tube inserted into the liquid outlet hole (11) of the chip serves as the negative power supply, and the power electrode (7) serves as the positive power supply. The signals detected by the two detection electrodes are connected to a differential amplifier circuit to perform noise reduction, filtering, and amplification processing on the signals. A USB DAQ 580-i is then used as a signal acquisition module to collect and analyze the detection signals.

[0043] Take 20 μL of 8 μm diameter particles and put them into 50 ml of 0.5% potassium chloride conductive solution. Shake the prepared particle solution evenly. First, use a precision injection pump to extract 10 mL of particle-free potassium chloride solution and inject the solution into the microfluidic chip at a rate of 10 mL / min for exhaust and lubrication. Then use the same steps to inject the prepared particle solution. The particle solution enters the inlet reservoir (4) through the liquid inlet hole (3), and then passes through the impurity filtering micro-column array (41) to filter out large-volume impurities. The filtered particle solution enters the curved inertial focusing channel (5). The Dean flow generated by the particles in the curved inertial focusing channel (5) and the secondary flow generated by the semicircular focusing boss (6) work together to form a high degree of focus. The focused particles flow through the upper and lower electrode holes (84, 94) for counting and detection. A DC voltage of 2V (decomposition voltage of the alloy electrode) is passed from the power supply electrode (7). The modules of the counting platform are connected to the corresponding working voltage to start collecting detection data. After passing through the detection electrode, the particle solution flows into the outlet reservoir (10) and flows out through the liquid outlet (11).

[0044] The capillary restriction valve detection electrode fabrication method described in this example only requires basic experimental equipment such as a hot plate, a vacuum dryer, and a syringe pump, significantly reducing the cost of fabricating the detection electrode. It also eliminates the steps required to design and configure a corresponding intelligent injection system, simplifying the electrode fabrication process.

[0045] The electrical signal obtained in the embodiment is as follows Figure 10As shown, Figure 10 The signal detected within 0.2 seconds obtained in the experiment, of which the larger amplitude signal is caused by the target particles flowing through the detection hole, the shorter signal is caused by the impurity particles, and the large number of small amplitude signals in the figure are noise signals. The noise signal is taken as 0.7 millivolts, and we get Figure 11 , then take Figure 11 The larger amplitude signal is finally obtained Figure 12 The particle signal shown is shown. Clearly, the chip possesses efficient particle counting detection capabilities. After traversing the data, the noise signal is taken as 0.002V. Furthermore, the signal-to-noise ratio of the data detected by the chip measured in the embodiment is 6, which is a high signal-to-noise ratio. This indicates that the chip is capable of detecting particles with a diameter of 15 microns, and the detection throughput can reach 30 particles per second, far exceeding the detection throughput of 3 particles per 100 seconds achieved by Gao Lingeng. This solves the clogging problem that occurs when the capillary restriction valve detection chip detects higher concentrations of particles, confirming the ability of the focused flow channel to optimize this structure.

[0046] Comparative Example

[0047] The focusing channel of this scheme adopts a constant width design, and the secondary flow vector distribution diagram generated in the end cross section is as follows: Figure 9 As shown, Figure 8 It can be seen from the comparison that the secondary flow intensity of the present invention is greater and the focusing effect is better.

Claims

1. A microfluidic impedance detection chip for particle counting, characterized in that: The invention comprises an upper substrate (1) and a lower substrate (2) which are bonded to each other, wherein the lower substrate (2) is provided with an inlet liquid reservoir (4), a curved inertial focusing flow channel (5) and a particle detection component, and the two ends of the curved inertial focusing flow channel (5) are respectively connected to the inlet liquid reservoir (4) and the particle detection component; the curved inertial focusing flow channel (5) is surrounded by an outer boundary (51) and an inner boundary (52) to form a flow channel, wherein the outer boundary (51) is an arc shape and the inner boundary (52) is a The Archimedean spiral is formed so that the width of the curved inertial focusing channel (5) decreases gradually along the direction from the inlet liquid reservoir (4) to the particle detection component; a plurality of evenly distributed focusing bosses (6) are provided on one side of the inner boundary (52) close to the outer boundary (51); the upper substrate (1) is provided with a liquid inlet hole (3) and a liquid outlet hole (11), the liquid inlet hole (3) is connected to the inlet liquid reservoir (4), and the liquid outlet hole (11) is connected to the particle detection component.

2. The microfluidic impedance detection chip for particle counting according to claim 1, characterized in that: The particle detection assembly comprises an outlet liquid reservoir (10) connected to an end of the curved inertial focusing channel (5) away from the inlet liquid reservoir (4), and the outlet liquid reservoir (10) is connected to a power supply electrode (7), an upper detection electrode (8) and a lower detection electrode (9).

3. The microfluidic impedance detection chip for particle counting according to claim 2, characterized in that: The power electrode (7) includes a power electrode liquid reservoir (71) and a power electrode filling channel (72), wherein the power electrode filling channel (72) is used to connect the power electrode liquid reservoir (71) and the outlet liquid reservoir (10), and the upper substrate (1) is provided with a power electrode liquid inlet hole (12), wherein the power electrode liquid inlet hole (12) is connected to the power electrode liquid reservoir (71).

4. The microfluidic impedance detection chip for particle counting according to claim 2, characterized in that: The upper detection electrode (8) comprises an upper detection electrode liquid reservoir (81) and an upper detection electrode filling channel (82), wherein the upper detection electrode filling channel (82) is used to connect the upper detection electrode liquid reservoir (81) and the outlet liquid reservoir (10), and the upper substrate (1) is provided with an upper detection electrode liquid inlet hole (13), wherein the upper detection electrode liquid inlet hole (13) is connected to the upper detection electrode liquid reservoir (81).

5. The microfluidic impedance detection chip for particle counting according to claim 2, characterized in that: The lower detection electrode (9) comprises a lower detection electrode liquid reservoir (91) and a lower detection electrode filling channel (92), wherein the lower detection electrode filling channel (92) is used to connect the lower detection electrode liquid reservoir (91) and the outlet liquid reservoir (10), and the upper substrate (1) is provided with a lower detection electrode liquid inlet hole (14), wherein the lower detection electrode liquid inlet hole (14) is connected to the lower detection electrode liquid reservoir (91).

6. The microfluidic impedance detection chip for particle counting according to claim 1, characterized in that: An impurity filtering micro-column array (41) is provided in the inlet liquid storage tank (4).

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