A microfluidic dielectrophoresis chip and its manufacturing method and application in liquid metal particle sorting

Abstract

The present invention relates to a microfluidic dielectrophoresis chip and a manufacturing method thereof and an application in liquid metal particle sorting. The microfluidic dielectrophoresis chip comprises a microelectrode and a microchannel. The microelectrode comprises a glass substrate and two metal chromium electrodes located on the glass substrate. Each metal chromium electrode comprises a wave segment and a pin segment connected to the end of the wave segment. The wave segments of the two metal chromium electrodes are parallel to each other, and the pin segments of the two metal chromium electrodes are mutually divergent. The microchannel is a PMDS cover sheet with a concave channel on the lower surface. The microchannel is bonded and packaged with the microelectrode through PMDS, and the wave segments of the two metal chromium electrodes are both located in the concave channel of the microchannel. The present invention realizes the particle sorting and capturing functions of liquid metal particles through dielectrophoresis microfluidic technology, and its sorting efficiency reaches more than 95%, which can broaden the application of dielectrophoresis microfluidic technology in biomarker detection and cell sorting.

Description

Technical Field

[0001] The invention relates to the field of sorting chips, in particular to a microfluidic dielectrophoresis chip and a manufacturing method thereof and an application thereof in liquid metal particle sorting. Background Art

[0002] In recent decades, microfluidics has attracted extensive attention due to its advantages such as miniaturization, portability, integration, automation, low cost, high throughput and simple operation. It is an interdisciplinary field that integrates perspectives from chemistry, physics, life sciences, microelectronics, materials science, computer science, etc., and has been applied to in vitro diagnostics (IVD), liquid biopsy, environmental and biochemical analysis, single cell analysis, nucleic acid analysis, etc. More specifically, conventional analytical analysis steps such as mixing, separation, enrichment, manipulation, sorting detection, synthesis and cell culture are realized in small chips and microfluidic systems. The ability of microfluidics to quickly and efficiently separate cells usually relies on external force fields such as optics, electronics, magnetic fields or acoustics. BAW operation based on acoustic flow and radiation force has a wide range of application prospects, and its advantages include versatility, biocompatibility, precision, flexibility, compactness and cost-effectiveness, as well as easy integration with other microfluidic technologies. To date, a wide range of particles with a size range from millimeters to millimeters in various fluid media (such as air, whole blood or sputum) have been successfully controlled using this technology.

[0003] Current status of microfluidic technology development; Microfluidic dielectrophoresis technology has developed rapidly in recent years. In 1999, Morgan et al. designed a polynomial electrode to capture and classify tobacco mosaic virus (TMV) and herpes simplex virus (HSV) type I; Li et al. proposed a wavy microfluidic dielectrophoresis chip in 2012, which achieved the separation of polystyrene microparticles and yeast cells under the application of different DC voltages; Pilloni et al. proposed a new design in 2016, using individually addressable planar and three-dimensional carbon microelectrodes to manipulate particles in three-dimensional space through dielectrophoretic forces.

[0004] L. Yu, C. Iliescu and others designed a discontinuous flow separation chip model. The shape and size of the microchannel of this model can change, resulting in changes in the electric field distribution in the channel and forming a non-uniform electric field. The working process of the chip is as follows: two mixed samples A and B are passed into the microchannel. Due to the effect of the non-uniform electric field, the dielectric responses of different samples are different, resulting in the samples gathering at different positions in the channel. At this time, pressure is applied at one end of the channel, and the A particles gathered in the middle of the channel will flow with the air pressure and flow out of the channel first. After all the A particles in the channel flow out, the electrical signal in the channel is disconnected. The B particles in the groove eventually flow out with the air flow due to the lack of the dielectrophoretic force, thus achieving the separation of the two mixed samples in time. In order to achieve the separation operation of particles of different sizes, Yusukawa and others improved the cross-finger electrode in 2009. The design concept of this structure is relatively novel. Through the relationship that the dielectrophoretic force is proportional to the cube of the particle diameter, a microelectrode is designed. When particles of different sizes pass through the channel, they are gradually distributed at different positions of the electrode structure due to the effect of forces of different magnitudes. The right picture shows that when a mixed solution of two particles with diameters of 10μm and 3μm is injected into the channel, both particles are affected by the negative dielectrophoretic force and are mixed together through the flow channel. After a period of time, it can be seen in the outlet area that there are only particles with a diameter of 3μm between the two narrow electrodes, while particles with a diameter of 10μm are between the wide and narrow electrodes. This is because the particles with larger diameters in the channel are gradually transferred to the designated area under the strong electric field at the wide electrode. The research on separation using dielectrophoresis has entered a period of rapid development since the 1990s. Many research groups in the world have realized the operation of particle separation using dielectrophoresis, such as Morgan et al., who separated polystyrene microspheres of different sizes, and Yang et al., who performed differential analysis of white blood cells; domestic institutions such as Tsinghua University and Southeast University have also conducted in-depth research in this field.

[0005] Yang Chaoyu et al. were inspired by the new concept of a three-dimensional fluid control platform based on a unit structure and proposed a view of tissue engineering based on the construction of a vascular network of cell fluids. Red blood cells [6-8 μm in diameter] are one of the key indicators in clinical medicine. They are separated from whole blood and have important practical significance because many biological targets span the same size range. For example, Huang et al. separated exosomes with high blood cell removal rates from whole blood by integrating high-frequency (39.4 MHz) interdigital transducers (IDTs). They demonstrated an acoustic fluidic platform that can encode droplets through (FIDTs) with an optimal driving frequency of 96.25 MHz. Lee et al. optimized the design of a high-frequency (394 MHz) IDT and underlying electronics to isolate nano- and microscale vesicles from cell culture media with high separation yield and resolution. In addition, Huang et al. designed a simple, low-cast nature, and open fluidic chamber platform that utilizes phase modulation between two low-frequency [4-6 kHz] piezoelectric transducers to achieve dynamic particle concentration and particle vortex translation, achieving single-direction operation of only one vortex.

[0006] Heavy metal inhalation caused by daily eating habits or environmental factors or heavy metal accumulation caused by food will cause metal particles in human blood, and the accumulation of metal ions in human blood will cause chronic poisoning in the human body, which is the primary factor leading to human disease and premature aging. At the same time, in many fields, from food and water safety to environmental monitoring and clinical analysis, they are closely related to human health. Therefore, purifying harmful particles such as metal particles in blood or drinking water can prevent and treat diseases, which is of great significance to human health. The metal particles accumulated in human blood include liquid metal particles. At present, no one has used microfluidic technology to manipulate liquid metal particles and achieve their sorting in blood. Summary of the invention

[0007] The present invention aims to provide a microfluidic dielectrophoresis chip and a manufacturing method thereof and application in liquid metal particle sorting, so as to realize particle sorting and capturing functions of liquid metal particles through dielectrophoresis microfluidic technology.

[0008] In order to solve the above technical problems, the specific scheme adopted by the present invention is: a microfluidic dielectrophoresis chip, including a microelectrode and a microchannel, the microelectrode including a glass substrate and two metal chromium electrodes located on the glass substrate, each metal chromium electrode including a wavy segment and a pin segment connected to the end of the wavy segment, the wavy segments of the two metal chromium electrodes are parallel to each other, and the pin segments of the two metal chromium electrodes are away from each other; the microchannel is a PMDS cover sheet with a concave channel on the lower surface, the microchannel is bonded and packaged with the microelectrode through PMDS, and the wavy segments of the two metal chromium electrodes are both located in the concave channel of the microchannel.

[0009] As a further optimization of the above technical solution, the width of the wave segment is 100 μm, the spacing between the wave segments of the two metal chromium electrodes is 100 μm, and the width of the sorting area formed by the two wave segments is 850 μm.

[0010] As a further optimization of the above technical solution, the microchannel includes a channel body, two inlets located at one end of the channel body and an outlet located at the other end of the channel body, the channel body includes an inlet section, a sorting section and an outlet section connected in sequence, one end of the sorting section is connected to the two inlets through the two inlet sections, and the other end of the sorting section is connected to the outlet through an outlet section.

[0011] As a further optimization of the above technical solution, the depth of the concave channel is 0.5 mm, the width of the sorting section is 2 mm, and the widths of the inlet section and the outlet section are both 1 mm.

[0012] A method for manufacturing a microfluidic dielectrophoresis chip comprises the following steps

[0013] S1: Fabrication of microelectrodes: Metal chromium electrodes were fabricated on glass substrates by photolithography and wet etching to form microelectrodes;

[0014] S2: Making a microchannel: taking another glass substrate, enclosing a containing cavity on the glass substrate, fixing a microchannel positive mold in the containing cavity, pouring PDMS into the containing cavity and performing a curing treatment to form a PMDS cover sheet; after taking the PMDS cover sheet out of the containing cavity, taking out the positive mold, thus forming a microchannel, wherein the position of the positive mold is a concave channel;

[0015] S3: Bonding and packaging of microelectrodes and microchannels: PMDS is applied to the lower surface of the PMDS cover sheet and covered on the glass substrate where the metal chromium electrode is located, so that the wavy section of the metal chromium electrode is located in the concave channel. After curing treatment, the microfluidic dielectrophoresis chip is obtained.

[0016] As a further optimization of the above technical solution, in step S2, the accommodating cavity is surrounded by a glass substrate and four glass slides, and the four glass slides are connected end to end and are all bonded to the glass substrate.

[0017] As a further optimization of the above technical solution, in step S2, the male mold includes a slab and protrusions fixed at both ends of the slab and used to form the inlet and outlet of the microchannel respectively.

[0018] As a further optimization of the above technical solution, in step S2, the height of the PDMS poured in the accommodating cavity is not higher than the height of the protrusion.

[0019] As a further optimization of the above technical solution, in step S2, the curing treatment is specifically: heating at an ambient temperature of 80°C for 30 minutes.

[0020] A microfluidic dielectrophoresis chip is used in liquid metal particle sorting. First, a solution to be sorted containing liquid metal particles is introduced into a concave channel from an inlet of a microchannel. After the concave channel is filled with the solution to be sorted, an ultrasonic power amplifier connected to the microfluidic dielectrophoresis chip and a function signal generator connected to the ultrasonic power amplifier are turned on. The parameters of the function signal generator are adjusted and a non-uniform electric field is provided to the microfluidic dielectrophoresis chip after amplification by the ultrasonic power amplifier. The liquid metal particles are captured by a metal chromium electrode under the action of the non-uniform electric field. Then, the inlet for the solution to be sorted is closed, and a medium solution is introduced from another inlet of the microchannel. Meanwhile, the ultrasonic power amplifier is disconnected. After the liquid metal particles lose the capture of the metal chromium electrode, they flow out of the concave channel with the medium solution to achieve sorting.

[0021] Compared with the prior art, the beneficial effects of the present invention are as follows: the present invention realizes the sorting and capturing functions of liquid metal particles through dielectrophoresis microfluidic technology, and its sorting efficiency reaches more than 95%, which can broaden the application of dielectrophoresis microfluidic technology in cell sorting, biomedical engineering, medical equipment and other fields. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is a schematic diagram of the structure of a microfluidic dielectrophoresis chip;

[0023] Figure 2 is an enlarged view of the microelectrode;

[0024] Figure 3 Flow chart for the fabrication of microelectrodes;

[0025] Figure 4 Schematic diagram of the overall structure of the microelectrode;

[0026] Figure 5 It is the microchannel size diagram;

[0027] Figure 6 is a schematic diagram of the structure of the accommodating cavity;

[0028] Figure 7 It is a structural schematic diagram of the positive mold;

[0029] Figure 8 is a schematic diagram of the male mold after being fixed on the accommodating cavity;

[0030] Fig. 9 This is a schematic diagram of the force on the particles in the microchannel during the sorting of the EGaln-red blood cell mixed solution;

[0031] Fig.10This is a schematic diagram of the distribution of eutectic gallium indium and red blood cells before power is applied during the sorting of EGaln-red blood cell mixed solution;

[0032] Fig.11 This is a schematic diagram of the distribution of eutectic gallium and indium after power is applied during the sorting of EGaln-red blood cell mixed solution;

[0033] Fig.12 This is a schematic diagram of the distribution of red blood cells after power is applied during the sorting of the EGaln-red blood cell mixed solution;

[0034] Fig.13 This is the distribution diagram of the ratio of eutectic gallium indium and red blood cells in the solution before and after power-on when the EGaln-red blood cell mixed solution was sorted by ImageJ software;

[0035] Fig.14 This is a real picture of the sorting effect of single eutectic gallium-indium particles;

[0036] Fig.15 It is a mixed solution sample of eutectic gallium-indium and red blood cells;

[0037] Fig.16 Schematic diagram of a sorting operation system equipped with a microfluidic dielectrophoresis chip. DETAILED DESCRIPTION

[0038] Example 1

[0039] like Figure 1 , Figure 2 and Figure 4 As shown, this embodiment is a microfluidic dielectrophoresis chip, including a microelectrode and a microchannel, wherein the microelectrode includes a glass substrate and two metal chromium electrodes located on the glass substrate.

[0040] The glass substrate is made of quartz, and each metal chromium electrode includes a wave segment and a pin segment connected to the end of the wave segment. The wave segments of the two metal chromium electrodes are parallel to each other, and the pin segments of the two metal chromium electrodes are away from each other; the width of the wave segment is 100μm, the spacing between the wave segments of the two metal chromium electrodes is 100μm, and the width of the sorting area formed by the two wave segments is 850μm. In this embodiment, the pin segments of the two metal chromium electrodes are parallel to each other and are located on both sides of the width direction of the sorting area, and the thickness of the chromium layer of the metal chromium electrode is about 100 nanometers.

[0041] Since the wave shape of the wave segment can also be called "W-type", the metal chromium electrode with a wave segment in this embodiment is also called a "W-type electrode". Compared with point electrodes and straight electrodes, the W-type electrode has a higher capture efficiency.

[0042] like Figure 1 , Figure 5As shown, the microchannel is a PMDS cover sheet with a concave channel on the lower surface, the microchannel is bonded and packaged with the microelectrode through PMDS, and the wave segments of the two metal chromium electrodes are both located in the concave channel of the microchannel. The wave segment has the same length direction as the concave channel and is located at the center of the concave channel.

[0043] The microchannel includes a channel body, two inlets at one end of the channel body and an outlet at the other end of the channel body, the channel body includes an inlet section, a sorting section and an outlet section connected in sequence, one end of the sorting section is connected to the two inlets through the two inlet sections respectively, and the other end of the sorting section is connected to the outlet through an outlet section. The depth of the concave channel is 0.5 mm, the width of the sorting section is 2 mm, the width of the inlet section and the outlet section are both 1 mm, and the total length of the concave channel is 35 mm.

[0044] The two inlet sections are an upper inlet section and a lower inlet section, the inlet end of the upper inlet section is set as an upper inlet, and the inlet end of the lower inlet section is set as a lower inlet, and the outlet end of the upper inlet section and the outlet end of the lower inlet section are connected to the sorting section after intersection, wherein the lower inlet section, the sorting section and the outlet section are coaxially distributed, and the angle between the upper inlet section and the lower inlet section is an acute angle.

[0045] Example 2

[0046] This embodiment is a method for manufacturing the microfluidic dielectrophoresis chip described in embodiment 1, comprising the following steps:

[0047] S1: Fabrication of microelectrodes: Metal chromium electrodes were fabricated on glass substrates by photolithography and wet etching to form microelectrodes;

[0048] Specifically, the process flow is as follows: Figure 3 As shown, the production method is mainly divided into the following steps:

[0049] a. Cleaning: First, clean the glass substrate in an ultrasonic cleaner with deionized water. The glass substrate is made of quartz. After cleaning with deionized water, place the quartz glass substrate in a prepared acid solution for pickling. After the pickling is completed, use deionized water to clean it. The cleaning work seems simple, but it is not. If it is not cleaned thoroughly, the microelectrode produced will contain impurities, which will affect the results.

[0050] b. Electroplating: Fix the cleaned quartz glass substrate in the electroplating tank of the electroplating studio, set the parameters of the electroplating gun, and electroplate a chromium layer of about 100 nanometers.

[0051] c. SU-8 photoresist spin coating: Place the quartz glass substrate after electroplating the chrome layer on the spin coating platform, and use the spin coater to spin coat a layer of SU-8 photoresist. The SU-8 photoresist after spin coating is evenly distributed on the surface of the glass substrate. Place the quartz glass substrate after spin coating in a vacuum drying oven at 80°C, heat for 30 minutes, and wait for the solvent to evaporate to increase the adhesion between the SU-8 photoresist and the glass substrate.

[0052] d. Exposure: Exposure is the most important step. If the exposure is not good, it will affect the accuracy of microelectrode production. Put the quartz glass substrate after spin coating into the UV exposure machine, turn on the UV exposure machine, set the exposure time to 5 seconds, and perform exposure.

[0053] e. Development: Place the quartz glass substrate after exposure treatment in a configured developer for development to remove unnecessary photoresist.

[0054] f. Wet etching: After removing the excess photoresist, wet etching is required to leave the required electrode chromium layer and etch away the unnecessary chromium layer. Place the quartz glass substrate after development in the prepared acid solution and etch for 3 minutes.

[0055] g. Cleaning: The quartz glass substrate after wet etching needs to be cleaned with deionized water.

[0056] h. Glue removal: This operation is to remove the glue on the electrode to expose the electrode.

[0057] i. Cleaning: Clean the chromium electrode.

[0058] k. Laser cutting and packaging: At this point, the production of the metal chromium electrode is complete. The quartz glass substrate is cut to the required size and then packaged to prevent dust from falling into it. Figure 4 shown.

[0059] S2: Making a microchannel: Take another glass substrate, enclose a containing cavity on the glass substrate, fix the microchannel positive mold in the containing cavity, pour PDMS into the containing cavity and perform a curing treatment to form a PMDS cover sheet; after taking the PMDS cover sheet out of the containing cavity, take out the positive mold, and then form a microchannel, wherein the position of the positive mold is a concave channel.

[0060] Before making the microchannel, it is necessary to design the microchannel first. Combined with the design of the microelectrode, the depth of the concave channel in the microchannel cannot be too deep, otherwise the particles will be suspended in the solution and too far away from the non-uniform electric field, and the dielectrophoretic force will be very small; the width of the microchannel cannot be too wide, otherwise a large number of particles will not flow over the microelectrode, but will flow along the channel wall from both ends of the microelectrode. The designed channel size is as follows: Figure 5As shown, the depth of the concave channel is 0.5 mm, the width of the sorting section is 2 mm, the widths of the inlet section and the outlet section are both 1 mm, and the total length of the concave channel is 35 mm.

[0061] The specific steps include:

[0062] 1) Preparation of the accommodating cavity: Use four Chuanfan brand glass slides with a size of 25 mm × 75 mm. First, fix one glass slide to a large glass substrate using 502 glue, and then use 502 glue to connect the four glass slides end to end and fix them to the large glass substrate. After the fixation is completed, a square cavity is left in the middle as the accommodating cavity for making the microchannel. The square cavity made is as follows: Figure 6 As shown, it should be noted that the accommodating cavity may be a cavity of other shapes besides a square cavity.

[0063] 2) Positive mold production: In order to save costs and control the thickness of the microchannel, a 0.5mm thick acrylic plate is used to make the positive mold. First, use AutoCAD2020 version software to draw the graphics of the positive mold, and output the drawn figure as a .wmf format file. Then use CorelLASER software to open this file, set the cutting speed to 2mm / s, the number of cuts to 3 times, and the current parameter to 8A. After the parameters are set, place the 0.5mm acrylic plate on the cutting platform of the laser engraving machine, start cutting, and wait for the cutting to be completed. The final positive mold is as follows Figure 7 shown.

[0064] 3) Fabrication of microchannels: First, use deionized water to clean the male mold and the square cavity. After cleaning, set three protrusions on the male mold. Specifically, use double-sided tape to fix three small cylinders to the inlet and outlet of the male mold, and then use 502 glue to fix the male mold with cylinders in the center of the square cavity.

[0065] After the positive mold is fixed, start to configure PDMS. Cut off half of the disposable paper cup and place it on the analytical balance. After peeling, add PDMS first until the analytical balance shows a weight of 10g, then add 1g of curing agent, mix the configured PDMS evenly, and pour it into the square cavity. The height of the PDMS poured in the cavity should not be higher than the height of the protrusion. After standing for a period of time, put it in a vacuum drying oven and evacuate for 30 minutes. After evacuating, heat it in a microwave oven at 80°C for 30 minutes to promote the curing of PDMS. After the PDMS is heated and cured, take it out of the PMDS cover square cavity formed, and then take out the positive mold. The position of the positive mold is the concave channel, and the position of the protrusion forms the entrance and exit of the concave channel, that is, the entrance and exit of the microchannel, thereby completing the production of the microchannel. The microchannel when the positive mold is not taken out is as shown Figure 8 shown.

[0066] S3: Bonding and packaging of microelectrodes and microchannels: Apply a layer of PMDS on the lower surface of the PMDS cover sheet and quickly cover it on the quartz glass substrate where the metal chromium electrode is located, so that the wavy section of the metal chromium electrode is located in the concave channel, and then heat it in a microwave oven at 80°C for 30 minutes for curing to obtain a microfluidic dielectrophoresis chip. The microfluidic dielectrophoresis chip bonded with PDMS is both firm and does not leak solution. The chip after bonding and packaging is as follows Figure 1 shown.

[0067] Example 3

[0068] This embodiment is an application of the above-mentioned microfluidic dielectrophoresis chip in liquid metal particle sorting. The microfluidic dielectrophoresis chip particle sorting system can be used to manipulate liquid metal particles to achieve the sorting of liquid metal particles and whole blood, and can also sort individual liquid metal particles.

[0069] When the microfluidic dielectrophoresis chip is used in the liquid metal particle sorting process, the microfluidic dielectrophoresis chip needs to be set in a sorting operation system, and the sorting operation system also includes a signal transmission device, a particle transmission device, and a data acquisition device.

[0070] The signal transmission device includes a function signal generator and an ultrasonic power amplifier, which provide a signal of a specific frequency, a specific waveform, and a specific voltage for the microfluidic dielectrophoresis chip. The frequency, peak-to-peak value, waveform, and phase are pre-set, and the function signal generator sends the set signal, which is amplified by the ultrasonic power amplifier to provide a non-uniform electric field for the microfluidic dielectrophoresis chip.

[0071] The particle delivery device includes a microinjection pump to provide particles to the microfluidic dielectrophoresis chip. In order to control the speed of the particles in the microfluidic dielectrophoresis chip, a microinjection pump is selected, which can not only meet the demand for particle movement speed but also control the movement of the syringe in both directions (i.e., forward propulsion and reverse extraction).

[0072] The microfluidic dielectrophoresis chip is the core of the entire sorting operating system and the key to particle sorting. After receiving the signal amplified by the ultrasonic power amplifier, the chip generates a non-uniform electric field. The particles are polarized in the non-uniform electric field to generate dielectric force, which causes them to deviate under the action of dielectric force, thereby achieving particle sorting.

[0073] The data acquisition device includes a microscope and a computer, which are used to observe and record the operation phenomena. A Leica microscope imported from Germany is selected and connected to the microscope through a computer. The operation phenomena can be observed through the computer display, and the movement trajectory and sorting status of the particles can be recorded by photographing or recording at any time.

[0074] The application process of microfluidic dielectrophoresis chip in liquid metal particle sorting is as follows:

[0075] First, a solution to be separated containing liquid metal particles is introduced into the inner concave channel from an inlet of the microchannel. Specifically, the lower inlet of the microchannel is kept closed, and the solution to be separated containing liquid metal particles is introduced into the inner concave channel from the upper inlet.

[0076] After the solution to be sorted fills the concave channel, the ultrasonic power amplifier connected to the microfluidic dielectrophoresis chip and the function signal generator connected to the ultrasonic power amplifier are turned on, the parameters of the function signal generator are adjusted and a non-uniform electric field is provided to the microfluidic dielectrophoresis chip after amplification by the ultrasonic power amplifier, and the liquid metal particles are captured by the metal chromium electrode under the action of the non-uniform electric field;

[0077] Then close the inlet for the solution to be sorted, and introduce the medium solution from another inlet of the microchannel, that is, close the upper inlet, open the lower inlet, and introduce the medium solution from the lower inlet. At the same time, disconnect the ultrasonic power amplifier. After the liquid metal particles lose the capture of the metal chromium electrode, they flow out of the concave channel with the medium solution to achieve sorting.

[0078] Eutectic gallium indium particles (EGaln) are a type of liquid metal particles. Therefore, taking eutectic gallium indium particles as an example, the sample solution (i.e., the solution to be sorted) is sorted through a microfluidic dielectrophoresis chip, wherein the sample solution containing liquid metal particles can be a sample solution containing only eutectic gallium indium particles, or it can be a mixed sample solution containing eutectic gallium indium particles and red blood cells.

[0079] 1. The operation process of continuous sorting of single eutectic gallium-indium particles by dielectrophoresis chip is as follows:

[0080] 1) Take out the packaged microfluidic dielectrophoresis chip (DEC chip for short), check whether the DEC chip is damaged, and use a soldering iron to solder the two copper wires to the two pins of the DEC chip. Finally, use a multimeter to check whether the welding is qualified. The DEC chip can be used only after the inspection is qualified.

[0081] 2) Turn on the power of the operating system and check whether the instruments in the operating system can work normally. After checking, stick a layer of double-sided tape around the bottom of the welded dielectrophoresis chip to fix the dielectrophoresis chip under the microscope. Adjust the microscope light source to transmitted light and the magnification to five times. Adjust the focal length, first coarsely adjust the focus to find the dielectrophoresis chip sorting area, then fine-focus to clearly see the dielectrophoresis chip sorting area on the computer monitor, and finally adjust the light intensity to avoid the light brightness being too high or too low, which will affect the shooting quality. It should be noted that the dielectrophoresis chip sorting area refers to the area where the concave channel of the microchannel and the wave section of the metal chromium electrode are located.

[0082] 3) Connect a 5 ml syringe to the infusion tube, then shake the eutectic gallium indium microparticle sample solution, quickly draw the syringe in the opposite direction, so that the eutectic gallium indium microparticles are in the infusion tube, and hang the infusion tube vertically. The end of the infusion tube away from the syringe is connected to the inlet of the microinjection pump. The outlet of the microinjection pump is connected to the upper inlet of the dielectrophoresis chip through a rubber hose, that is, the upper inlet is connected to the eutectic gallium indium microparticle suspension. Take another 5 ml syringe, draw 5 ml of the medium solution, and connect the syringe to the inlet of another microinjection pump through the infusion tube. The outlet of the microinjection pump is connected to the lower inlet of the dielectrophoresis chip through a rubber hose, that is, the lower inlet is connected to the medium solution. It is worth noting here that because the eutectic gallium indium suspension precipitates too quickly, most of the eutectic gallium indium microspheres are usually precipitated after 1 minute of mixing and shaking, so the eutectic gallium indium microsphere sample solution cannot be directly placed in a syringe, otherwise the eutectic gallium indium microspheres will precipitate, and it will be impossible to transport the eutectic gallium indium microspheres to the dielectrophoresis chip, and continuous sorting cannot be completed. Therefore, it is chosen to store the eutectic gallium indium microsphere sample solution in an infusion tube, and transport the microspheres through the infusion tube, thereby reducing the impact caused by the precipitation of the eutectic gallium indium microspheres.

[0083] 4) After the microparticle delivery device is connected, set the flow rate or flow rate, turn on the microinjection pump for delivering the sample solution, turn off the microinjection pump for delivering the medium solution, and slowly push the syringe connected to the sample solution infusion tube to fill the rubber hose with the eutectic gallium indium microsphere sample solution first, and then fill the dielectrophoresis chip. At the same time, pay attention to whether bubbles are generated. The presence of bubbles will affect the movement direction and speed of the eutectic gallium indium microspheres, so the pushing speed of the syringe should not be too fast. If bubbles are generated, reduce the speed of the microinjection pump, slowly pass the solution, and expel the bubbles. If the bubbles cannot be expelled, the dielectrophoresis chip can only be removed, cleaned, and then dried to operate again.

[0084] 5) When the sample solution of the eutectic gallium indium microspheres is observed to completely fill the dielectrophoresis chip and no bubbles are generated, adjust the flow rate of the microinjection pump, and first observe and photograph the movement trajectory of the eutectic gallium indium microspheres without power. After the observation and photography are completed, connect the output end of the ultrasonic power amplifier to the copper wire on the dielectrophoresis chip, adjust the voltage, frequency and other parameters, observe and photograph the movement of the eutectic gallium indium microspheres in the dielectrophoresis chip sorting area, and use ImageJ and other software to perform statistical analysis on the particles.

[0085] 6) The eutectic gallium indium microparticles are captured by the microelectrode under the action of the positive dielectrophoretic force, the microinjection pump for delivering the sample solution is turned off, the microinjection pump for delivering the medium solution is turned on, and the connection between the ultrasonic power amplifier and the dielectrophoresis chip is disconnected at the same time, so that the metal chromium electrode loses its adsorption and capture effect, and the medium solution enters the sorting area from the lower inlet to flush out the eutectic gallium indium microparticles captured by the metal chromium electrode, thereby realizing the collection of the sorted eutectic gallium indium microparticles.

[0086] By repeating steps 3) to 6) with different dielectric solutions, the effect of different dielectric solution conductivity on the continuous sorting efficiency of eutectic gallium-indium particles can be studied. It should be noted that when turning off the ultrasonic power amplifier, the connection between the ultrasonic power amplifier and the dielectrophoresis chip should be disconnected first to avoid the generation of many bubbles and damage to the electrodes due to the drastic change in voltage when the ultrasonic power amplifier is turned off.

[0087] After all operations are completed, the dielectrophoresis chip is cleaned with deionized water. After the chip is cleaned under a microscope, it is dried and packaged for next use. Finally, all instruments are turned off and the operating platform is cleaned.

[0088] The results of sorting single eutectic gallium indium particles are as follows:

[0089] The optimal parameters are a frequency of 100 kHz, a suspension flow rate of 50 μl / min, a voltage of 20 V, and a suspension conductivity of 7 μS / m. At this time, the separation efficiency of eutectic gallium-indium particles is as high as 99%. Fig.14 shown.

[0090] 2. Sorting and capturing of EGaln-red blood cell mixed solution;

[0091] The sample solution in this test process is a mixed sample solution. The configuration method of the mixed sample solution is as follows: the particles used are mainly eutectic gallium indium with a radius of 15um and red blood cells with a radius of 5um.

[0092] Preparation of eutectic gallium-indium suspension solution: Take 100 ml of deionized water and add it into a beaker. Use a pipette to add 10 ul of eutectic gallium-indium solution. After diluting it 100 times, shake it for 1 minute to make the eutectic gallium-indium in the diluted solution evenly distributed to obtain the desired eutectic gallium-indium ball suspension solution.

[0093] Preparation of red blood cell suspension solution: first prepare 100 ml of normal saline, then use a blood collection needle to take a drop of blood from the human body, add it to the prepared normal saline to dilute it, and oscillate for 1 minute to make the red blood cells evenly distributed in the diluted solution to obtain the required red blood cell suspension solution.

[0094] Preparation of mixed sample solution: Use a pipette to extract 10μl of eutectic gallium indium microparticles, place them in a 5ml centrifuge tube, add PBS phosphate buffered saline to 5ml, then use a pipette to extract 50μl of red blood cells, place them in another 5ml centrifuge tube, and also add PBS phosphate buffered saline to 5ml. After repeated mixing at different proportions, a mixed sample solution with a ratio of eutectic gallium indium microparticles to red blood cells of 1:100 is finally prepared. Fig.15 shown.

[0095] The operation process of the dielectrophoresis chip for sorting eutectic gallium-indium microparticles and red blood cells is as follows:

[0096] 1) Take out the packaged and bonded dielectrophoresis chip and check whether the dielectrophoresis chip is damaged. After checking, use a soldering iron to solder the two copper wires to the two pins of the dielectrophoresis chip respectively, and then use a multimeter to check whether the welding is qualified. The dielectrophoresis chip can be used only after the inspection is qualified.

[0097] 2) Turn on the power of the operating system and check whether all instruments in the operating system can work normally. After checking, fix the dielectrophoresis chip under the microscope and adjust the microscope so that the sorting area of ​​the dielectrophoresis chip can be clearly seen on the computer monitor.

[0098] 3) Connect a 5 ml syringe to the infusion tube, then shake the mixed sample solution to mix it evenly, quickly draw the syringe in the reverse direction, the mixed sample solution is in the infusion tube, the infusion tube is vertically suspended, the end of the infusion tube away from the syringe is connected to the inlet of the micro-injection pump, the outlet of the micro-injection pump is connected to the upper inlet of the dielectrophoresis chip through a rubber hose, that is, the upper inlet is connected to the mixed sample solution. Take another 5 ml syringe, draw 5 ml of PBS phosphate buffered saline solution as the medium solution, connect the syringe to the inlet of another micro-injection pump through the infusion tube, the outlet of the micro-injection pump is connected to the lower inlet of the dielectrophoresis chip through a rubber hose, and the lower inlet is connected to the PBS phosphate buffered saline solution.

[0099] 4) After the particle delivery device is connected, first turn on the microinjection pump that delivers the mixed sample solution, then slowly push the syringe to allow the mixed sample solution to fill the rubber hose first to expel bubbles, set the mixed sample solution flow rate to 50μl / min, and allow it to slowly fill the dielectrophoresis chip. At the same time, pay attention to whether there are bubbles generated. The presence of bubbles will affect the movement direction and speed of the eutectic gallium indium microspheres. If bubbles are generated, use tweezers to gently press the dielectrophoresis chip position corresponding to the bubble above until the bubble is discharged or becomes very small and basically does not affect the operation.

[0100] 5) When it is observed that the eutectic gallium indium microspheres completely fill the chip and no bubbles are generated, observe and photograph the movement trajectory of the eutectic gallium indium microspheres without power. After the observation and photography are completed, connect the output end of the ultrasonic power amplifier to the copper wire on the dielectrophoresis chip, adjust the peak-to-peak value of the function signal generator to 5V, the frequency to 100kHz, the waveform to a sine wave, and the phase to 0. Once the output channel switch is turned on, the signal is transmitted to the ultrasonic power amplifier, and then the ultrasonic power amplifier is adjusted to make the output voltage 15V. Observe and photograph the movement of the eutectic gallium indium microspheres and red blood cells in the dielectrophoresis chip sorting area, and use ImageJ and other software to perform statistical analysis on the particles.

[0101] 6) Use a microfluidic dielectrophoresis chip to sort red blood cells and eutectic gallium indium microparticles. The eutectic gallium indium microparticles are captured by the microelectrode under the action of the positive dielectrophoretic force, while the dielectrophoretic force on red blood cells is much smaller than the fluid dynamics of the solution in the dielectrophoresis chip. Under the action of the fluid dynamics, the red blood cells flow out of the outlet with the solution; turn off the microinjection pump that delivers the mixed sample solution, turn on the microinjection pump that delivers the medium solution, and disconnect the ultrasonic power amplifier from the dielectrophoresis chip at the same time, so that the metal chromium electrode loses its adsorption and capture effect. The flow rate of the medium solution is 50μl / min, and the medium solution uses PBS phosphate buffered saline with a conductivity of 15μS / min. The medium solution enters the sorting area from the lower inlet to flush out the eutectic gallium indium microparticles captured by the metal chromium electrode, thereby collecting the sorted eutectic gallium indium microparticles and achieving the purpose of sorting.

[0102] One thing that needs to be explained is that the connection between the ultrasonic power amplifier and the dielectrophoresis chip should be disconnected first, and then the ultrasonic power amplifier should be turned off. Finally, the dielectrophoresis chip should be cleaned with deionized water. After the chip is cleaned under a microscope, it should be dried and packaged for the next use. Then all instruments should be turned off and the platform should be cleaned.

[0103] <Analysis of the force acting on particles in the dielectrophoresis chip>

[0104] In terms of EGaln sorting and capture: the force analysis of particles in the microfluidic dielectrophoresis chip is as follows:

[0105] The forces acting on particles in the dielectrophoresis chip include not only the dielectrophoretic force, but also buoyancy, gravity, fluid dynamics, fluid resistance, intermolecular forces, etc. Compared with the dielectrophoretic force and fluid dynamics, the other forces are too small, so the dielectrophoretic force F acting on the particles is mainly considered. DEP and fluid dynamics F l The schematic diagram of the force acting on the particle is shown in Figure 9, which can be used to represent the resultant force F acting on the particle in the dielectrophoresis chip:

[0106]

[0107] When no AC electric field and flow rate are applied (i.e., E = 0, V = 0), the particles are in a state of equilibrium in the dielectrophoresis chip and do not move. When an AC electric field and flow rate are applied to the dielectrophoresis chip, the particles are in a state of equilibrium in the dielectrophoresis chip and do not move. DEP and fluid dynamics F l The particle will move under the action of , so the motion equation of the particle in the dielectrophoresis chip can be expressed as follows:

[0108]

[0109] In the above formula: m m is the mass of the particle, in kg; v m is the movement speed of particles in the dielectrophoresis chip, in m / s; It is the resultant force acting on the particle in the dielectrophoresis chip, in N.

[0110] Since the particles used are spherical particles, their mass is:

[0111]

[0112] In the above formula, r is the radius of the particle, in m; ρ m is the density of the particle, in kg / m 3 .

[0113] In the microfluidic dielectrophoresis chip, the particle displacement x m (t) can be expressed as:

[0114]

[0115] Without considering the acceleration process of particles in the microfluidic field, only considering the uniform motion of particles, then in the x direction:

[0116] F DEP sinθ=F l

[0117] F l =6πμηr(v m-x -v m )

[0118] In the z direction:

[0119] F DEP cosθ=F drag

[0120] F drag =6πηrv m-z

[0121] From this, the movement speed of the particles in the microfluidic dielectrophoresis chip can be obtained:

[0122]

[0123]

[0124] v m-x represents the movement speed of the particles in the x - direction in the dielectrophoresis chip, and v m-z represents the movement speed of the particles in the z - direction in the dielectrophoresis chip. η represents the pseudorapidity of the particles, and θ represents the angle between the resultant force F and the hydrodynamic force F l .

[0125] <Sorting Results of EGaIn - Red Blood Cell Mixed Solution>

[0126] In the present invention, a wavy micro - electrode is adopted. The flow rate of the micro - injection pump is adjusted to 150 μm / s, and the function signal generator is adjusted to 0.04 V and 500 kHz. Then, a mixed sample solution of eutectic gallium - indium and red blood cells is added to a 5 - ml syringe and connected to the upper inlet. After all the devices are debugged, the micro - injection pump for transporting the sample solution is turned on, and the micro - injection pump for transporting the medium solution is turned off. The movement state of the particles in the microscope is observed on the computer, as Fig.10 .

[0127] From Fig.10 , it can be seen that before the power is applied, the eutectic gallium - indium and red blood cells around the electrode are scattered, but Fig.11 it can be clearly seen that after the power is applied, the eutectic gallium - indium around the electrode is captured by the metal chromium electrode, while the red blood cells directly pass through the metal chromium electrode, and Fig.12 there are a large number of red blood cells around the channel outlet. Then, we use ImageJ software to count and statistically analyze the three pictures in Fig.10 , Fig.11 , Fig.12 , and obtain the proportion of eutectic gallium - indium and red blood cells in the solution as Fig.13 .

[0128] Through Fig.13 , it can be seen that there is only 10% of eutectic gallium - indium in the solution before the power is applied. After the power is applied, the eutectic gallium - indium is captured by the electrode, while the red blood cells can still pass through. Therefore, the proportion of eutectic gallium - indium in the solution increases, and the proportion of red blood cells in the solution at the outlet is as high as 100%, achieving the effect of separating eutectic gallium - indium and red blood cells.

Claims

1. A microfluidic dielectrophoresis chip, characterized in that: The invention comprises a microelectrode and a microchannel. The microelectrode comprises a glass substrate and two metal chromium electrodes located on the glass substrate. Each metal chromium electrode comprises a wavy segment and a pin segment connected to the end of the wavy segment. The wavy segments of the two metal chromium electrodes are parallel to each other, and the pin segments of the two metal chromium electrodes are away from each other. The microchannel is a PMDS cover sheet with a concave channel on the lower surface. The microchannel is bonded and packaged with the microelectrode through PMDS. The wavy segments of the two metal chromium electrodes are both located in the concave channel of the microchannel. The microchannel comprises a channel body, two inlets located at one end of the channel body and an outlet located at the other end of the channel body. The channel body comprises an inlet segment, a sorting segment and an outlet segment which are connected in sequence. One end of the sorting segment is connected to the two inlets through the two inlet segments respectively, and the other end of the sorting segment is connected to the outlet through an outlet segment.

2. The microfluidic dielectrophoresis chip according to claim 1, characterized in that: The width of the wave segment is 100 μm, the spacing between the wave segments of the two metal chromium electrodes is 100 μm, and the width of the separation area formed by the two wave segments is 850 μm.

3. The microfluidic dielectrophoresis chip according to claim 1, characterized in that: The depth of the concave channel is 0.5 mm, the width of the sorting section is 2 mm, and the widths of the inlet section and the outlet section are both 1 mm.

4. The method for manufacturing a microfluidic dielectrophoresis chip according to claim 1, characterized in that: The method comprises the following steps S1: making a microelectrode: making a metal chromium electrode on a glass substrate by photolithography and wet etching to form a microelectrode; S2: Making a microchannel: taking another glass substrate, enclosing a containing cavity on the glass substrate, fixing a microchannel positive mold in the containing cavity, pouring PDMS into the containing cavity and performing a curing treatment to form a PMDS cover sheet; after taking the PMDS cover sheet out of the containing cavity, taking out the positive mold, thus forming a microchannel, wherein the position of the positive mold is a concave channel; S3: Bonding and packaging of microelectrodes and microchannels: PMDS is applied to the lower surface of the PMDS cover sheet and covered on the glass substrate where the metal chromium electrode is located, so that the wavy section of the metal chromium electrode is located in the concave channel. After curing treatment, the microfluidic dielectrophoresis chip is obtained.

5. The method for manufacturing a microfluidic dielectrophoresis chip according to claim 4, characterized in that: In step S2, the accommodating cavity is surrounded by a glass substrate and four glass slides, and the four glass slides are connected end to end and bonded to the glass substrate.

6. The method for manufacturing a microfluidic dielectrophoresis chip according to claim 4, characterized in that: In step S2, the male mold includes a slab and protrusions fixed at both ends of the slab and used to form an inlet and an outlet of the microchannel respectively.

7. The method for manufacturing a microfluidic dielectrophoresis chip according to claim 6, characterized in that: In step S2, the height of the PDMS poured in the accommodating cavity is not higher than the height of the protrusion.

8. Use of the microfluidic dielectrophoresis chip according to claim 1 in liquid metal particle sorting.

9. The use of a microfluidic dielectrophoresis chip in liquid metal particle sorting according to claim 8, characterized in that: First, a solution to be sorted containing liquid metal particles is introduced into the microchannel from an inlet of the microchannel. After the solution to be sorted fills the concave channel, an ultrasonic power amplifier connected to the microfluidic dielectrophoresis chip and a function signal generator connected to the ultrasonic power amplifier are turned on. The parameters of the function signal generator are adjusted and a non-uniform electric field is provided to the microfluidic dielectrophoresis chip after amplification by the ultrasonic power amplifier. The liquid metal particles are captured by the metal chromium electrode under the action of the non-uniform electric field. Then, the inlet for the solution to be sorted is closed, and a medium solution is introduced from another inlet of the microchannel. At the same time, the ultrasonic power amplifier is disconnected. After the liquid metal particles lose the capture of the metal chromium electrode, they flow out of the concave channel with the medium solution to achieve sorting.

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