CN112457978A - Blood cell separation chip based on bulk acoustic wave and application - Google Patents
Blood cell separation chip based on bulk acoustic wave and application Download PDFInfo
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Abstract
The invention belongs to the technical field of cell chips, and particularly relates to a blood cell separation chip based on bulk acoustic waves, a system and application thereof.A blood cell separation plate is provided with a blood cell separation channel, and the blood cell separation channel is composed of a double-spiral channel; the blood cell separation channel is provided with three ports, namely a channel inlet, a channel outlet I and a channel outlet II, wherein the channel outlet I and the channel outlet II are separated from one channel; the chip is also provided with a sound wave generator which is positioned below the chip bottom plate; the chip cover plate, the blood cell separation plate, the chip bottom plate and the sound wave generator are integrally attached from top to bottom. The invention meets the requirement of high-throughput and instant separation of blood cells, combines the bulk acoustic wave technology with the double helix chip, realizes the effective separation of white blood cells in a blood sample to obtain the white blood cell-free blood, and has high white blood cell separation efficiency and good blood cell morphology.
Description
Technical Field
The invention belongs to the technical field of cell chips, and particularly relates to a blood cell separation chip based on bulk acoustic waves and application thereof.
Background
With the development of medical technology, the application range of clinical blood transfusion and component blood transfusion is gradually expanded, blood cell separation is an important link for blood treatment before blood transfusion, after a donor donates whole blood, transfusion can cause leukocyte-mediated transfusion reaction and related virus transmission due to input of allogeneic blood and preparations thereof, and leukocyte-removal transfusion can ensure safer transfusion and has important significance for clinical treatment of some diseases.
The main methods adopted for preparing blood components by blood cell separation at present are centrifugation, filtration, magnetic separation, antigen-antibody adsorption and the like, and the methods need bulky centrifugation equipment, expensive reagent materials or complex manufacturing procedures, are difficult to miniaturize and achieve high flux, and are difficult to meet the current blood cell separation requirements. And has the technical problems of complex flow, low separation efficiency, high cost and the like.
"microfluidics" refers to fluids used in experiments on the order of milliliters, micro-liters down to nanoliters or picoliters. The micro-fluidic chip developed by biotechnology is manufactured on silicon and glass by using a conventional plane processing technology (photoetching, corrosion and the like), or is formed by a soft photoetching micro-processing method for printing and forming a micro-structure on an organic material. The micro-fluidic chip system has the advantages of light volume, small amount of used samples and reagents, high reaction speed, capability of massively parallel processing, disposability and the like. The micro-acoustic fluid system power intensity and frequency are kept in a biocompatible range, the technology of manipulating particles in fluid by utilizing ultrasonic standing waves is well applied, and the acoustic manipulation technology ensures no damage of a living body sample due to non-contact, so that the micro-acoustic fluid system power intensity and frequency are a hotspot in the field of clinical research and application. Essentially, the acoustic treatment process does not change the properties of cells, and is now increasingly applied in the fields of tumor cell separation, exosome separation, and blood analysis and counting.
Chinese patent CN201310703508.3 specification describes a surface acoustic wave microfluidic chip for tumor cell separation, which comprises a substrate and a cover plate adhered on the substrate and provided with a microchannel; the micro-channel comprises a middle channel with an inlet and an outlet, wherein cell sap injection channels connected with the inlet are arranged on two sides of the inlet, and cell collection channels connected with the outlet are arranged on two sides of the outlet; two pairs of interdigital converters with consistent frequency and sound intensity are sequentially deposited on the substrates on two sides of the middle channel between the injection channel and the collection channel, and the distance from the second pair of interdigital converters to the center of the middle channel is 1/4 surface wave wavelengths greater than the distance from the first pair of interdigital converters to the center of the middle channel; and small cell collecting channels are symmetrically arranged on two sides of the middle channel between the two pairs of interdigital converters. The invention has the advantages that the standing wave type surface acoustic wave technology is adopted, and the tumor cells (CTCs) are separated efficiently, economically and pollution-free. However, the separation is difficult because of the small number of circulating tumor cells in the blood.
Blood cell separation refers to the separation of red blood cells, white blood cells and platelets in blood, which are used to prepare blood components that can be returned to the body if they are of high purity. Tumor cell isolation refers to the isolation of circulating tumor cells from the blood of a cancer patient for subsequent analysis of cancer information. Both are greatly different. The concentration of blood cells in blood is very high, the content of white blood cells is very low, the efficient separation of the white blood cells from the whole blood in the microfluidic chip still has great challenges, the integration and miniaturization difficulty is high, and the chip blockage and the separation efficiency are low.
Leukocytes are generally spherical, and most leukocytes have a diameter greater than 10 microns and a larger size. Other blood cells, such as red blood cells, have an average diameter of about 6 microns, are 2 microns thick, and are relatively small in size. It is pointed out that separation is performed by using blood cell size, for example, a cross-flow cell separation chip described in "BioMEMS microfluidic chip for blood sample pretreatment" is formed by anodic bonding of glass (cover plate) and a silicon wafer (substrate) processed by MEMS technology, and the silicon substrate includes three flow cells and a microchannel. The glass cover plate comprises sample inlets, red blood cell sample outlets and white blood cell sample outlets which respectively correspond to the three flow cells of the siliceous base plate. The micro-channel on the silicon substrate is divided into three separated micro-channels by two parallel micro-column arrays, and the micro-column arrays are composed of micro-cylinders. The cross-flow filtration means that water flows parallel to the surface of the filtering microstructure and forms larger shearing force on the filtering surface, so that the blocking substances of the filtering microstructure are always in a suspension state, the accumulation of the blocking substances in the filtering microstructure is reduced, and the blockage of a chip can be effectively avoided. However, the technical problem of low efficiency of leukocyte separation still exists.
Disclosure of Invention
In order to solve the technical problems, the invention provides a blood cell separation chip based on bulk acoustic waves, which overcomes the defects of complex flow, low separation efficiency, high manufacturing cost and the like of the existing blood cell separation technology, meets the blood cell separation requirement of high-throughput and instant separation, combines the bulk acoustic wave technology with a double helix chip, realizes the effective separation of white blood cells in a blood sample to obtain the white blood cell-free blood, and has high white blood cell separation efficiency and good blood cell morphology; saving cost, high biological safety and simple preparation.
The invention solves the technical problem, provides a blood cell separation chip based on bulk acoustic waves, which comprises a blood sample inlet, a white blood cell outlet, other blood cell outlets, a chip cover plate, a blood cell separation plate and a chip bottom plate, wherein the blood sample inlet, the white blood cell outlet and the other blood cell outlets are arranged on the chip cover plate, the blood cell separation plate is arranged between the chip cover plate and the chip bottom plate, and the blood cell separation chip is characterized in that: the blood cell separation plate is provided with a blood cell separation channel which is composed of a double-spiral channel; the blood cell separation channel is provided with three ports, namely a channel inlet, a channel outlet I and a channel outlet II, wherein the channel outlet I and the channel outlet II are separated from one channel; the chip is also provided with a sound wave generator which is positioned below the chip bottom plate; the chip cover plate, the blood cell separation plate, the chip bottom plate and the sound wave generator are integrally attached from top to bottom.
The blood cell separation channel is a double-spiral channel, and the positive spiral and the negative spiral are connected by an S-shaped channel.
The total spiral turns of the blood cell separation channel are 8-14 turns; the spiral pitch in the optimization scheme is 480-520 microns. The spiral channel pitch (fluid does not pass) refers to the distance between two channels and the channel (fluid) width refers to the width of one channel.
The cross section of the blood cell separation channel is rectangular, and the height of the blood cell separation channel is half wavelength of sound waves generated by the sound wave generator; the width of the blood cell separation channel in the optimization scheme is 300-800 microns; in a further optimization scheme, the width of the blood cell separation channel is 500-700 microns; in a more preferred embodiment, the blood cell separation channel has a width of 500 μm.
The sound wave generator and the chip base plate are assembled together in a mode of smearing adhesive, and the used adhesive is ultrasonic couplant, epoxy resin and the like.
The sound wave generator is made of piezoelectric materials such as piezoelectric ceramic pieces or piezoelectric crystals; in the optimization scheme, the sound wave generator is a piezoelectric ceramic piece. The invention aims to form standing waves in the channel, and the piezoelectric ceramic plate has good piezoelectric effect and large electromechanical coupling coefficient.
The input power of the piezoelectric ceramic plate is 10-50dbm, and the input power of the piezoelectric ceramic plate under the optimized scheme is 30 dbm.
The working frequency of the piezoelectric ceramic piece is 500kHz-5MHz, the height of the blood cell separation channel is half wavelength of sound waves generated by the piezoelectric ceramic piece, and h is 3000/f.
In the optimization scheme, the working frequency of the piezoelectric ceramic plate is 1.5MHz, the height of the blood cell separation channel is half wavelength of sound wave generated by the piezoelectric ceramic plate, h is 3000/f is 200 micrometers, and the width of the blood cell separation channel is 500 micrometers.
If a stable standing wave field is to be formed in the intermediate blood cell separation channel, the height of the intermediate blood cell separation channel is equal to a half wavelength of the sound wave generated by the piezoelectric ceramic plate (channel height h is 3000/f). The wavelength of the sound wave corresponds to the frequency of the piezoelectric ceramic piece, according to the formula λ ═ c/f (c is the speed of light, λ is the wavelength of light wave (in air) when propagating in vacuum, and f is the frequency, and the unit is hertz), the propagation speed c of the sound wave in blood is approximately 1500m/s, and the working frequency of the piezoelectric ceramic piece is 500kHz-5 MHz.
The material of the blood cell separation channel plate is polymethyl methacrylate (PMMA), Polydimethylsiloxane (PDMS) or glass. The chip cover plate and the chip bottom plate are made of glass materials, so that the biocompatibility is good, and the materials can be obtained more conveniently and saved. The upper and lower parts of the invention are packaged by glass as sound wave resonant cavities, and the piezoelectric ceramic piece is used as a sound wave generator.
The invention comprises a system of a blood cell separation chip based on bulk acoustic waves, wherein a blood sample inlet is connected with a sample injector through a pvc infusion tube, the sample injector is fixedly connected with an injection pump, and a leukocyte outlet and other blood cell outlets are respectively connected with other devices; the injection pump injects the blood sample into the blood cell separation channel at a constant speed, and the injection flow rate is 5-30 mL/min; in the optimization scheme, the injection pump injects the blood sample into the blood cell separation channel at a constant speed, the injection flow rate is 15mL/min, different blood cells in the sample are separated in the chip, and finally the blood cells flow out from different outlets of the chip. The blood sample is whole blood or white membrane.
The bulk acoustic wave chip is used for removing white blood cells in a blood sample; the bulk acoustic wave chip is used for preparing blood components.
The technical principle of the invention is as follows: when the blood sample flows in the channel in a laminar flow state, the velocity of the fluid in the channel is distributed in a parabolic shape from the center to the outer wall, namely, the velocity is the fastest at the center and is smaller as the velocity is closer to the outer wall. Due to the fluid velocity gradient, the flow velocity of the fluid at the top and bottom of the blood cells is different, and the blood cells are subjected to shear-induced lift by the fluid. When the particles move to the channel wall under the action of the shear-induced force, the particles and the channel wall interact, the outer wall generates a force which prevents the particles from being close to the wall surface, the force is called wall surface induced lift force, and the wall surface induced lift force and the shear-induced lift force are called inertial lift force collectively.
The blood cells in the curved channel are subjected to the drag action of dean vortex in addition to the inertial lift force. The dean vortex is perpendicular to the flowing direction, and two swirling flows with opposite rotating directions are distributed up and down along the center of the channel. The presence of dean vortex introduces dean drag which carries blood cells in a circular motion along the channel center to the upper and lower walls, respectively. Thus, in addition to moving forward with the fluid, the blood cells also move in a vortex within the channel from the center of the channel to the upper and lower walls.
A sound wave generator is arranged at the lower part of the chip, taking a piezoelectric ceramic plate as an example, the piezoelectric ceramic plate is connected with a power amplifier through a lead, the piezoelectric ceramic plate generates sound waves through electro-acoustic conversion, sound wave signals are adjusted through a controllable signal generator and the power amplifier, the sound waves are transmitted into a middle blood cell separation channel and are reflected by an upper glass surface, reflected waves with the same frequency and amplitude as incident waves and in the opposite propagation direction are generated (the sound waves enter the channel and are reflected by the upper glass surface, the propagation directions of the waves are opposite), and the two lines of waves are superposed to form a stable standing wave sound field in the channel. Under the action of a standing wave sound field, most blood cells are subjected to the action of acoustic radiation force to generate displacement and are finally stabilized at node positions, and due to the action of standing wave fields at all positions in the channel, the node positions are at the same height to form node surfaces, most blood cells in the blood sample move to the node surfaces, and eddy current existing in the channel in the vertical direction is restrained.
The blood cells stabilized at the nodal plane only consider the stress in the horizontal direction, and the blood cells are subjected to the inertial lifting force F in the channelLAnd dean drag force FDThe blood cells reach a balance point, the induced lifting force and the dean force of the blood cells with different sizes in the same fluid are different, so that the balance positions of the blood cells are different, as shown in fig. 3, most of the blood cells are gathered on the nodal planes with the same height, the blood cells with different sizes are gradually gathered into lines along with the high-speed flow of the blood cells in the blood cell separation channel, the white blood cells flow out from an outlet closest to the inner wall of the channel due to the largest size of the white blood cells, and other blood cells are collected from another outlet, so that the white blood cells in the blood sample are removed. The blood cell separation channel is a double-spiral channel, and the schematic diagram is shown in fig. 4, wherein the forward spiral and the reverse spiral are connected by an S-shaped channel, so that the track of the blood cells in the channel after the blood cells are gathered is narrower.
According to the invention, a standing wave field is introduced into the blood cell separation channel, most blood cells are gathered at a certain height, and the blood cells with different sizes in the spiral channel are balanced differently, so that the blood sample can be efficiently removed from the white blood cells. The chip has high separation efficiency, the separation efficiency of the white blood cells reaches about 93 percent, the aim of removing the white blood cells in the blood sample is basically achieved, and the morphology of the blood cell cells is good.
The chip has the advantages of miniaturization and light weight, is simple in preparation process, is convenient for large-scale integrated preparation, and can be widely applied to the fields of life science, medicine and the like.
Drawings
The invention will be described in further detail with reference to the following drawings and detailed description:
FIG. 1 is a diagram showing a chip structure according to the present invention
FIG. 2 is a schematic diagram of the structure of the blood cell separation system based on bulk acoustic waves according to the present invention
FIG. 3 and FIG. 4 are schematic diagrams of the chip splitting structure of the present invention
FIG. 5 is a schematic diagram showing the equilibrium positions of blood cells of different sizes in the blood cell separation channel according to the present invention
FIG. 6 is a schematic view of the structure of a blood cell separation channel according to the present invention
FIG. 7 is a graph showing the effect of separating a sample of tunica albuginea using a blood cell separation chip based on bulk acoustic waves according to the present invention
Wherein the labels in the figure are specifically:
1. the blood sample inlet, 2, the white blood cell outlet, 3, the other blood cell outlet, 4, the chip upper glass cover plate, 5, the blood cell separation plate, 6, the lower chip bottom plate, 7, the piezoelectric ceramic plate, 8, the blood cell separation channel, 9, the channel inlet, 10, the channel outlet I, 11, the channel outlet II, 12, the pvc infusion tube, 13, the sample injector, 14, the injection pump, 15, the S-shaped channel
Detailed Description
The invention will be further illustrated with reference to specific embodiments:
example 1
A blood cell separation chip based on bulk acoustic waves is provided with a blood sample inlet, a leukocyte outlet, other blood cell outlets, a chip cover plate, a blood cell separation plate and a chip bottom plate, wherein the blood sample inlet, the leukocyte outlet and the other blood cell outlets are arranged on the chip cover plate; the blood cell separation channel is provided with three ports, namely a channel inlet, a channel outlet I and a channel outlet II, wherein the channel outlet I and the channel outlet II are separated from one channel; the piezoelectric ceramic plate is also arranged and is positioned below the chip bottom plate; the chip cover plate, the blood cell separation plate, the chip bottom plate and the piezoelectric ceramic plate are bonded into a whole from top to bottom. The blood cell separation channel is a double-helix channel, and the forward helix and the reverse helix are connected by an S-shaped channel.
The forward spiral and the reverse spiral are oppositely arranged, and if the channel inlet is connected with the forward spiral, the two channel outlets are connected with the reverse spiral. The total spiral turns of the blood cell separation channel are 8 turns; the pitch of the spirals was 480 microns.
The forward spiral and the reverse spiral are arranged oppositely, some blood cells under the forward spiral do not reach a focusing condition in the channel and are in a dispersed state in the channel, and when entering the reverse spiral through the S-shaped channel, most of the blood cells are more gathered on the track in the channel, so that different blood cells can flow out from the outlet. The S-shaped channel is designed to ensure that the track of the blood cells in the channel after the blood cells are gathered is narrower, and the positions of different blood cell tracks are different in the channel, so that the blood cells can be ensured to flow out from different outlets.
The piezoelectric ceramic plate and the chip base plate are assembled together by coating an adhesive, and the used adhesive is epoxy resin and the like. The blood cell separation channel plate material is polymethyl methacrylate (PMMA), and the chip cover plate and the chip bottom plate are glass materials.
The working frequency of the piezoelectric ceramic plate is 500kHz, the height of the blood cell separation channel is half wavelength of sound waves generated by the piezoelectric ceramic plate, the height of the blood cell separation channel is 600 micrometers, and the width of the blood cell separation channel is 300 micrometers. The input power of the piezoelectric ceramic plate is 20 dbm.
The bulk acoustic wave chip is used for removing white blood cells in a blood sample; or the bulk acoustic wave chip is used for preparing blood components.
Example 2
A blood cell separation chip based on bulk acoustic waves is provided with a blood sample inlet, a leukocyte outlet, other blood cell outlets, a chip cover plate, a blood cell separation plate and a chip bottom plate, wherein the blood sample inlet, the leukocyte outlet and the other blood cell outlets are arranged on the chip cover plate; the blood cell separation channel is provided with three ports, namely a channel inlet, a channel outlet I and a channel outlet II, wherein the channel outlet I and the channel outlet II are separated from one channel; the piezoelectric ceramic plate is also arranged and is positioned below the chip bottom plate; the chip cover plate, the blood cell separation plate, the chip bottom plate and the piezoelectric ceramic plate are bonded into a whole from top to bottom. The blood cell separation channel is a double-helix channel, and the forward helix and the reverse helix are connected by an S-shaped channel. The total spiral turns of the blood cell separation channel are 10 turns; the pitch of the helix was 500 microns.
The piezoelectric ceramic plate and the chip base plate are assembled together by smearing an adhesive, and the used adhesive is an ultrasonic coupling agent. The material of the blood cell separation channel plate is Polydimethylsiloxane (PDMS), and the material of the chip cover plate and the chip bottom plate is glass material.
The working frequency of the piezoelectric ceramic plate is 1.5MHz, the height of the blood cell separation channel is half wavelength of sound wave generated by the piezoelectric ceramic plate (h is 3000/f is 200 micrometers), the height of the blood cell separation channel is 200 micrometers, and the width of the blood cell separation channel is 500 micrometers. The input power of the piezoelectric ceramic plate is 30 dbm.
Example 3
The system comprises a blood cell separation chip based on bulk acoustic waves, wherein a blood sample inlet is connected with a sample injector through a pvc (polyvinyl chloride) infusion tube, the sample injector is connected with an injection pump, a blood sample is injected into a blood cell separation channel at a constant speed through the injection pump, and the injection flow rate is 5-30 mL/min; the blood sample is whole blood or white membrane.
The inlet of the blood sample is connected with a pvc infusion tube, the other end of the infusion tube is connected with a biological sample injector, the blood sample is injected into the chip at a certain constant speed through an injection pump, different blood cells in the sample are separated in the chip, and finally the blood cells flow out from different chip outlets.
The blood cell separation chip based on the bulk acoustic wave is provided with a blood sample inlet, a leukocyte outlet, other blood cell outlets, a chip cover plate, a blood cell separation plate and a chip bottom plate, wherein the blood sample inlet, the leukocyte outlet and the other blood cell outlets are arranged on the chip cover plate; the blood cell separation channel is provided with three ports, namely a channel inlet, a channel outlet I and a channel outlet II, wherein the channel outlet I and the channel outlet II are separated from one channel; the piezoelectric ceramic plate is also arranged and is positioned below the chip bottom plate; the chip cover plate, the blood cell separation plate, the chip bottom plate and the piezoelectric ceramic plate are bonded into a whole from top to bottom. The blood cell separation channel is a double-helix channel, and the forward helix and the reverse helix are connected by an S-shaped channel. The total spiral turns of the blood cell separation channel are 14 turns; the pitch of the spirals was 520 microns.
The piezoelectric ceramic plate and the chip base plate are assembled together by smearing an adhesive, and the used adhesive is an ultrasonic coupling agent. The blood cell separation channel plate is made of glass, and the chip cover plate and the chip bottom plate are made of glass.
The working frequency of the piezoelectric ceramic plate is 5MHz, the height of the blood cell separation channel is half wavelength of sound wave generated by the piezoelectric ceramic plate, the height of the blood cell separation channel is 60 micrometers, and the width of the blood cell separation channel is 700 micrometers. The input power of the piezoelectric ceramic plate is 10 dbm.
Example 4
Otherwise, as in example 1, the total number of spiral turns of the blood cell separation channel was 12; the pitch of the helix was 500 microns. The sound wave generator is made of piezoelectric crystal material, the blood cell separation channel has half of the wavelength, standing waves are formed in the blood cell separation channel, and the height of the blood cell separation channel is 200 micrometers. The width of the blood cell separation channel was 800 μm.
Wherein the injection pump injects the blood sample into the blood cell separation channel at a constant rate, and the injection flow rate is 15 mL/min. The bulk acoustic wave chip is applied to preparation of blood components.
Example 5
Otherwise as in example 1, the total number of turns of the spiral of the blood cell separation channel was 13 turns; the pitch of the spirals was 510 microns. The sound wave generator is made of a piezoelectric ceramic sheet material, the working frequency of the piezoelectric ceramic sheet is 800kHz, standing waves are formed in the blood cell separation channel, the height of the channel is half wavelength, the height of the channel is 937.5 micrometers, and the width of the blood cell separation channel is 600 micrometers. The input power of the piezoelectric ceramic plate is 50 dbm.
Wherein the injection pump injects the blood sample into the blood cell separation channel at a constant rate, and the injection flow rate is 15 mL/min. The bulk acoustic wave chip is applied to preparation of blood components.
Example 6
The steps of preparing the blood cell separation chip based on the bulk acoustic wave in the invention are as follows:
two glass sheets with the same size and thickness (about 300 microns) are cut, three round holes are cut in the upper glass cover plate by laser and are used as the inlet and the outlet of a blood sample, namely a blood sample inlet, a leukocyte outlet and other blood cell outlets, and the radius of the round hole of each outlet is 200-300 microns. The lowest layer is a chip bottom plate glass sheet which is complete glass and is used for packaging the cavity.
The middle layer between the two glass layers is a blood cell separation channel plate 5 made of Polydimethylsiloxane (PDMS), and the preparation process of the middle blood cell separation channel plate 5 comprises a series of processes of template design and preparation, photoetching, glue pouring, curing and punching. The height of the blood cell separation channel is adjustable according to the working frequency of the piezoelectric ceramic piece, the working frequency of the piezoelectric ceramic piece is 2MHz, the height of the corresponding middle layer separation channel 5 is equal to the half wavelength of sound waves generated by the piezoelectric ceramic piece, and therefore the height of the middle blood cell separation channel is 375 micrometers. The blood cell separation channel adopts a double-spiral channel, the total number of spiral turns is 9, and the width of the blood cell separation channel is 400 microns.
After the blood cell separation channel plate 5 is prepared, the lower glass bottom plate 6, the middle blood cell separation channel plate 5 and the upper glass cover plate 4 are tightly bonded together by adopting an oxygen plasma bonding method.
After the chip bonding is finished, the joints are respectively fixed at the three round holes in a glue bonding mode and respectively correspond to the blood sample inlet 1, the leucocyte outlet 2 and the other blood cell outlets 3. The connector may be a luer connector.
The connectors are fixed at the three round holes respectively in a glue bonding mode, so that the pvc pipe is inserted into the connectors, the connectors are fixed on the glass cover plate, and fluid flowing out of the blood cell separation channel enters the pvc pipe through the connectors to be collected. The dark portion of the port circle of fig. 6 is the location of the perforations where fluid flows inside the channel and where fluid flows inside the dark spiral channel. 123 in fig. 1 denotes a joint.
The three joints are respectively assembled with a pvc infusion tube for sample introduction and sample discharge, the other end of the infusion tube at the sample inlet is connected with an injection pump for injecting a blood sample into the chip at a constant speed, the flow rate is 15mL/min, the other end of the infusion tube at the sample outlet is connected with a leukocyte collecting tube, and the other end of the infusion tube connected to the sample outlet is connected with other blood cell collecting tubes.
After the blood cell separation channel plate 5 is bonded, an ultrasonic coupling agent layer is uniformly coated on the circular piezoelectric ceramic plate, the circular piezoelectric ceramic plate is attached to the chip bottom plate 6, and two copper wires are led out from two sides of the piezoelectric ceramic plate 7 to be used as signal input leads. By adjusting the signal generator and the power amplifier, different input powers, preferably 30dbm, are applied to the piezoceramic wafer 7.
Fig. 1 and 2 show the overall structure of the blood cell separation chip based on bulk acoustic waves, fig. 3 and 4 show the split structure of the chip, and fig. 6 shows the blood cell separation channel.
Test-separation of leukocytes from the white Membrane Using a bulk Acoustic wave-based blood cell separation chip
A white membrane sample is taken and blood cell separation is carried out by using the blood cell separation chip prepared by the embodiment, wherein the working frequency of the piezoelectric ceramic piece is 1.5MHz, the width of a channel of the blood cell separation chip is 500 micrometers, the height of the channel is 200 micrometers, the speed of injecting the sample into the blood cell separation channel is 15mL/min, and the input power of the piezoelectric ceramic piece is 30 dbm.
As shown in FIG. 7, when the diluted leucocyte sample passes through the blood cell separation chip of the present invention, the leucocytes of large size flow out from the inner channel, and most of the platelets of small size flow out from the outer channel.
Under an optical microscope, the blood cells with different sizes can be obviously separated better at the outlet, and the morphology of the blood cells is good. Through the conventional blood test of the sample collected at the outlet, the separation efficiency of the white blood cells reaches about 93 percent, and the aim of removing the white blood cells in the blood sample is basically fulfilled.
While the foregoing shows and describes the fundamental principles and principal features of the invention, together with the advantages thereof, the foregoing embodiments and description are illustrative only of the principles of the invention, and various changes and modifications can be made therein without departing from the spirit and scope of the invention, which will fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (10)
1. The utility model provides a blood cell separation chip based on bulk acoustic wave, including being equipped with blood sample entry (1), leucocyte export (2), other blood cell export (3), chip cover board (4), blood cell separation board (5) and chip bottom plate (6), blood sample entry (1), leucocyte export (2) and other blood cell export (3) are established on chip cover board (4), and its characterized in that is established between chip cover board (4) and chip bottom plate (6) in blood cell separation board (5): the blood cell separation plate (5) is provided with a blood cell separation channel (8), the blood cell separation channel (8) is composed of a double-spiral channel and is provided with three ports, namely a channel inlet, a channel outlet I and a channel outlet II, the channel outlet I and the channel outlet II are separated from one channel, the channel inlet is connected with a blood sample inlet (1), the channel outlet I is connected with a leukocyte outlet (2), and the channel outlet II is connected with other blood cell outlets (3); the sound wave generator is also arranged and is positioned below the chip bottom plate (6); the chip cover plate (4), the blood cell separation plate (5), the chip bottom plate (6) and the sound wave generator (7) are bonded into a whole from top to bottom.
2. The bulk acoustic wave-based blood cell separation chip according to claim 1, wherein: the blood cell separation channel is a double-spiral channel and is provided with a forward spiral and a reverse spiral, and the forward spiral and the reverse spiral are connected by an S-shaped channel.
3. The bulk acoustic wave-based blood cell separation chip according to claim 2, wherein: the total spiral turns of the blood cell separation channel (8) are 8-14 turns; the spiral channel spacing in the optimization scheme is 480- & 520 microns.
4. A bulk acoustic wave-based blood cell separation chip according to any one of claims 1 to 3, wherein: the cross section of the blood cell separation channel is rectangular, and the height of the blood cell separation channel is half wavelength of sound waves generated by the sound wave generator; the width of the blood cell separation channel in the optimization scheme is 300-800 microns; in a further optimization scheme, the width of the blood cell separation channel is 500-700 microns; in a more preferred embodiment, the blood cell separation channel has a width of 500 μm.
5. The bulk acoustic wave-based blood cell separation chip according to claim 1, wherein: the sound wave generator (7) is a piezoelectric ceramic piece or a piezoelectric crystal piezoelectric material; in the optimization scheme, the sound wave generator (7) is a piezoelectric ceramic piece.
6. The bulk acoustic wave-based blood cell separation chip according to claim 5, wherein: the input power of the piezoelectric ceramic plate (7) is 10-50dbm, and the input power of the piezoelectric ceramic plate (7) in the optimized scheme is 30 dbm.
7. The bulk acoustic wave-based blood cell separation chip according to claim 5, wherein: the working frequency of the piezoelectric ceramic piece (7) is 500kHz-5MHz, the height of the blood cell separation channel (8) is half wavelength of sound waves generated by the piezoelectric ceramic piece, and h is 3000/f; in the optimization scheme, the working frequency of the piezoelectric ceramic piece (7) is 1.5MHz, the height of the blood cell separation channel (8) is half wavelength of sound waves generated by the piezoelectric ceramic piece, h is 3000/f is 200 micrometers, and the height of the blood cell separation channel (8) is 200 micrometers.
8. A bulk acoustic wave-based blood cell separation chip according to any one of claims 1 to 3, wherein: the material of the blood cell separation channel plate (5) is polymethyl methacrylate (PMMA), Polydimethylsiloxane (PDMS) or glass; the chip cover plate (4) and the chip bottom plate (6) are made of glass materials.
9. A bulk acoustic wave based blood cell separation system according to claim 1, wherein: the blood sample inlet (1) is connected with a sample injector through a pvc infusion tube, the sample injector is fixedly connected with an injection pump, and the leukocyte outlet (2) and other blood cell outlets (3) are respectively connected with other devices; the injection pump injects the blood sample into the blood cell separation channel (8) at a constant speed, and the injection flow rate is 5-30 mL/min; in the optimization scheme, a blood sample is injected into a blood cell separation channel by an injection pump at a constant speed, and the injection flow rate is 15 mL/min; the blood sample is whole blood or white membrane.
10. The use of a bulk acoustic wave-based blood cell separation chip according to claim 1, wherein: the bulk acoustic wave chip is used for removing white blood cells in a blood sample; the bulk acoustic wave chip is used for preparing blood components.
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| CN118122404A (en) * | 2024-05-06 | 2024-06-04 | 北京理工大学 | Microfluidic chip |
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