Real-time high-flux portable separation device and method for multiple biological microparticles based on micro-fluidic chip

Abstract

The invention relates to a real-time high-flux portable separation device and method for multiple biological microparticles based on a micro-fluidic chip. The real-time high-flux portable separation device comprises a substrate (1) and a multi-particle automatic separation micro-fluidic chip (2) which is positioned above the substrate (1) and is based on a dielectrophoresis technology, a dielectrophoresis electrode (12) of the micro-fluidic chip is connected with the signal generator (4); a vibration exciter (5) is arranged above a compressible chamber (13) of the micro-fluidic chip; the particle collecting cavity (6), the microparticle sample pool (7) and the deionized water storage pool (8) are connected with an inlet and an outlet of the micro-fluidic chip through hoses (9). According to different parameters such as dielectric parameters, shapes and sizes of separation objects in the mixed samples, separation of three or more mixed samples is realized under the same frequency, and the device has the characteristics of automation, high throughput and portable real-time separation and has a good application prospect.

Description

Technical Field

[0001] The present invention belongs to the technical field of microfluidic chips, and particularly relates to a device and method for real-time high-throughput portable separation of various biological microparticles based on a microfluidic chip. Background Art

[0002] In the research in fields such as pathogen detection and drug development, microorganism screening is a key biomedical sensing technology in urgent need. In current research, firstly, for microorganisms themselves, there are a large number of microorganism species and huge amounts of data, and it is very difficult to separate microorganisms in complex matrices. Secondly, traditional microorganism screening methods often use methods such as plate culture and sequencing for "culturing first and then separating". This method has a large workload, requires a long time, needs professional personnel to operate, and cannot obtain the original information of microorganisms, let alone achieve on-site detection. Summary of the Invention

[0003] The present invention provides a device and method for real-time high-throughput portable separation of various biological microparticles based on a microfluidic chip. The device can achieve automated, high-throughput, and portable real-time separation of three or more types of bacterial microparticles, and has good application prospects.

[0004] The present invention provides a device for real-time high-throughput portable separation of various biological microparticles based on a microfluidic chip, including a substrate and a multi-particle automatic sorting microfluidic chip based on dielectrophoresis technology located above the substrate; the dielectrophoresis electrodes of the microfluidic chip are connected to a signal generator; an exciter is arranged above the pressable chamber of the microfluidic chip; the particle collection chamber, the microparticle sample pool, and the deionized water storage pool are all connected to the inlets and outlets of the microfluidic chip through hoses.

[0005] Further, the device is also provided with a micro-control unit, which serves as a control center to control the AC signal parameters of the signal generator and the working frequency and digital operation time of the exciter at the same time.

[0006] Further, the multi-particle automatic sorting microfluidic chip is composed of a quartz substrate and a PDMS layer; the microfluidic chip includes a microparticle sample inlet, a first deionized water inlet, a pressable chamber, a buffer chamber, a separation zone, a one-way valve, and a second deionized water inlet; dielectrophoresis electrodes are arranged on the quartz substrate.

[0007] Further, the separation zone is provided with a plurality of sub-separation channels and a main separation channel; the sub-separation channels are vertically distributed above the dielectrophoresis electrodes, and the main separation channel is distributed in the middle of the dielectrophoresis electrodes.

[0008] Further, a plurality of outlets are connected to the main separation channel.

[0009] The present invention also provides a method for using a real-time high-throughput portable separation device for multiple biological microparticles based on a microfluidic chip, including the following steps:

[0010] (1) First, add target microparticles and deionized water to the microparticle sample pool and the deionized water storage pool respectively to prepare for the separation experiment;

[0011] (2) Start the micro control unit and apply drive signals to the vibrator and the signal generator; among them, the vibrator acts on the pressable chamber periodically, that is, press - release. When the pressable chamber is in the pressed state, the one-way valve is opened, and the liquid in the pressable chamber is pumped out into the separation zone to achieve sample injection; when the pressable chamber is in the non-pressed state (i.e., the released state), the one-way valve will be in the closed state. At this time, the microparticle sample pool and the deionized water storage pool take the microparticle sample and deionized water, and the microparticle sample and deionized water will be sucked into the pressable chamber from the microparticle sample inlet and the second deionized water inlet; repeat the above pressing state to achieve automatic sample injection operation;

[0012] (3) When the microparticle sample flows into the separation zone, it first flows into the secondary separation channel. The electrical parameters are set by the signal generator so that one kind of particle in the mixed microparticle sample is adsorbed on the dielectrophoresis electrode under the action of positive dielectrophoresis force to achieve the separation of one kind of sample in the mixed microparticle sample; then, the remaining microparticle sample flows into the main separation channel and is squeezed on both sides of the channel close to the dielectrophoresis electrode under the action of the deionized water flowing in from the second deionized water inlet. At this time, the microparticles close to the dielectrophoresis electrode are separated into at least two other kinds of particles according to the differences in their own dielectric constants, particle sizes, and morphologies under the action of negative dielectrophoresis force. The separated particles flow out from the outlet and into the particle collection chamber respectively;

[0013] (4) Stop sample injection, pump deionized water from the first deionized water inlet to clean the separation zone and the secondary separation channel. After the deionized water all flows into the main separation channel, stop the signal generator. The first kind of particle adsorbed on the dielectrophoresis electrode detaches and flows out from the outlet and into the particle collection chamber;

[0014] (5) Repeat steps (1) to (4) to achieve real-time high-throughput portable separation of multiple biological microparticles.

[0015] Beneficial effects

[0016] (1) Through the matching design of the microchannel and the dielectrophoresis electrode, the present invention simultaneously realizes the discontinuous separation and continuous separation of microparticles under the same group of electrodes, and realizes the separation of three or more kinds of mixed samples at the same frequency according to the different dielectric properties, shapes, sizes, etc. of the separation objects in the mixed sample.

[0017] (2) The present invention is based on microfluidic technology, combines a pressable chamber, a one-way valve and a vibrator to replace the traditional injection pump, and realizes automatic sample injection of the device.

[0018] (3) The present invention combines the liquid stable flow splitting technology and the arrayed dielectrophoresis sorting technology to realize high-throughput detection of samples.

[0019] (4) The sample injection method of the present invention can realize programmable digital control and realize diversified operations of samples.

[0020] (5) The present invention can realize automatic, high-throughput and portable real-time separation of three or more kinds of bacterial microparticles, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 is a schematic structural diagram of the device of the present invention.

[0022] Figure 2 is a schematic structural diagram of a microfluidic chip for automatic sorting of multiple particles based on dielectrophoresis technology.

[0023] Figure 3 is a plan view of a microfluidic chip for automatic sorting of multiple particles based on dielectrophoresis technology; wherein, (a) top view; (b) bottom view.

[0024] Figure 4 is a schematic diagram of the separation area of a microfluidic chip for automatic sorting of multiple particles based on dielectrophoresis technology; wherein, (a) top view; (b) bottom view.

[0025] Figure 5 is a working flow chart of the device of the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0026] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. In addition, it should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the appended claims of this application.

[0027] Example 1

[0028] By Figure 1As shown in the figure, this embodiment provides a real-time high-throughput portable separation device for various biological microparticles based on a microfluidic chip, including a substrate 1 and a microfluidic chip 2 for automatic sorting of various particles based on dielectrophoresis technology located above the substrate 1; the dielectrophoresis electrodes 12 of the microfluidic chip are connected to a signal generator 4; an exciter 5 is arranged above the pressable chamber 13 of the microfluidic chip; a particle collection chamber 6, a microparticle sample pool 7, and a deionized water storage pool 8 are all connected to the inlet and outlet of the microfluidic chip through hoses 9. The device is also provided with a micro-control unit 3, which controls the AC signal parameters of the signal generator 4 and the operating frequency and digital operation time of the exciter 5 as a control center at the same time.

[0029] The signal generator 4 is connected to the dielectrophoresis electrodes 12, and the signal emitted is applied to the dielectrophoresis electrodes 12 to provide an AC signal for microparticle separation. The exciter 5 acts on the pressable chamber 13 of the microfluidic chip and is used to periodically and digitally press the pressable chamber 13.

[0030] As Figure 2 As shown in the figure, the microfluidic chip 2 for automatic sorting of various particles is composed of a quartz substrate 18 and a PDMS layer 19; the microfluidic chip 2 includes a microparticle sample inlet 10, a first deionized water inlet 11, a pressable chamber 13, a buffer chamber 14, a separation zone 16, a one-way valve 17, and a second deionized water inlet 15; the dielectrophoresis electrodes 12 are arranged on the quartz substrate 18.

[0031] When the exciter 5 acts on the pressable chamber 13, the pressable chamber 13 deforms towards the bottom of the channel, the one-way valve 17 closes, and the liquid in the pressable chamber 13 is pumped into the channel to achieve sample injection. When the exciter 5 moves away from the pressable chamber 13, the one-way valve 17 opens, the sample liquid enters the pressable chamber 13, and the pressable chamber 13 returns to its original shape. By periodically and digitally controlling the exciter 5, programmable sample injection control can be achieved. The buffer chamber 14 is provided to slow down the jitter of the fluid flow during sample injection in a buffered manner and make the fluid flow more stable.

[0032] As Figure 3 As shown in the figure, the dielectrophoresis electrodes 12 (interdigitated electrodes) are sputtered on the quartz substrate 18 by microfabrication technology, and the PDMS layer 19 containing microchannels is placed in the corresponding position through a bonding process.

[0033] As Figure 4 As shown in the figure, the separation zone 16 is provided with a plurality of secondary separation channels 20 and a main separation channel 21; the secondary separation channels 20 are vertically distributed above the dielectrophoresis electrodes 12, and the main separation channel 21 is distributed in the middle of the dielectrophoresis electrodes 12. A plurality of outlets 22 are connected to the main separation channel 21.

[0034] As Figure 5As shown, this embodiment also provides a method for using a real-time high-throughput portable separation device for multiple biological microparticles based on a microfluidic chip, including the following steps

[0035] (1) First, add target microparticles and deionized water to the microparticle sample pool 7 and the deionized water storage pool 8 respectively to prepare for the separation experiment;

[0036] (2) Start the micro-control unit 3 and apply drive signals to the vibrator 5 and the signal generator 4. Among them, the vibrator 5 acts on the pressable chamber 13 periodically, that is, press-release. When the pressable chamber 13 is in the pressed state, the one-way valve 17 is opened, and the liquid in the pressable chamber 13 is pumped out into the separation zone 16 to achieve sample injection. When the pressable chamber 13 is in the non-pressed state, the one-way valve 17 will be in the closed state. At this time, the microparticle sample pool 7 and the deionized water storage pool 8 take the microparticle sample and deionized water and will be sucked into the pressable chamber 13 from the microparticle sample inlet 10 and the second deionized water inlet 15. Repeat the above pressing state to achieve automatic sample injection operation;

[0037] (3) When the microparticle sample flows into the separation zone 16, it first flows into the secondary separation channel 20. The electrical parameters are set by the signal generator 4 so that one kind of particle in the mixed microparticle sample is adsorbed on the dielectrophoresis electrode 12 under the action of positive dielectrophoresis force to achieve the separation of one kind of sample in the mixed microparticle sample. Then, the remaining microparticle sample flows into the main separation channel 21 and is squeezed on both sides of the channel near the dielectrophoresis electrode 12 by the deionized water flowing in from the second deionized water inlet 15. At this time, the microparticles near the dielectrophoresis electrode 12 are separated into at least two other kinds of particles according to the differences in their own dielectric constants and particle sizes and shapes under the action of negative dielectrophoresis force. The separated particles flow out from the outlet 22 and flow into the particle collection chamber 6;

[0038] (4) Stop sample injection, pump deionized water from the first deionized water inlet 11 to clean the separation zone 16 and the secondary separation channel 20. Stop the signal generator after the deionized water all flows into the main separation channel 21. The first kind of particle adsorbed on the dielectrophoresis electrode 12 is detached and flows out from the outlet 22 and into the particle collection chamber 6;

[0039] (5) Repeat steps (1) to (4) to achieve real-time high-throughput portable separation of multiple biological microparticles.

Claims

1. A portable real-time high-throughput separation device for multiple biological microparticles based on a microfluidic chip, characterized by: The invention comprises a substrate (1) and a dielectrophoresis technology-based microfluidic chip (2) for automatically sorting multiple particles, located above the substrate (1); the dielectrophoresis electrode (12) of the microfluidic chip is connected to a signal generator (4); an exciter (5) is arranged above a pressable chamber (13) of the microfluidic chip; and a particle collection chamber (6), a microparticle sample pool (7) and a deionized water storage pool (8) are all connected to the inlet and outlet of the microfluidic chip via a hose (9).

2. The portable device for real-time high-throughput separation of multiple biological microparticles according to claim 1, characterized in that: The device is also provided with a micro control unit (3) which serves as a control center and simultaneously controls the AC signal parameters of the signal generator (4) and the operating frequency and digital operation time of the exciter (5).

3. The portable device for real-time high-throughput separation of multiple biological microparticles according to claim 1, characterized in that: The multi-particle automatic sorting microfluidic chip (2) is composed of a quartz substrate (18) and a PDMS layer (19); the microfluidic chip (2) comprises a microparticle sample inlet (10), a first deionized water inlet (11), a pressable chamber (13), a buffer chamber (14), a separation area (16), a one-way valve (17), and a second deionized water inlet (15); and a dielectrophoresis electrode (12) is arranged on the quartz substrate (18).

4. The portable device for real-time high-throughput separation of multiple biological microparticles according to claim 3, characterized in that: The separation zone (16) is provided with a plurality of secondary separation channels (20) and a main separation channel (21); the secondary separation channels (20) are vertically distributed above the dielectrophoresis electrode (12), and the main separation channel (21) is distributed in the middle of the dielectrophoresis electrode (12).

5. The portable device for real-time high-throughput separation of multiple biological microparticles according to claim 4, characterized in that: The main separation channel (21) is connected to a plurality of outlets (22).

6. A method for using the real-time high-throughput portable separation device for multiple biological microparticles based on a microfluidic chip according to any one of claims 1 to 5, comprising the following steps: (1) First, target microparticles and deionized water are added to the microparticle sample pool (7) and the deionized water storage pool (8) respectively to prepare for the separation experiment; (2) starting the microcontroller unit (3) and applying a driving signal to the exciter (5) and the signal generator (4); wherein, The exciter (5) acts on the depressible chamber (13) periodically, i.e., presses and releases. When the depressible chamber (13) is in a pressed state, the one-way valve (17) is opened, and the liquid in the depressible chamber (13) is pumped out to the separation zone (16) to achieve sample injection. When the depressible chamber (13) is in a non-pressed state, the one-way valve (17) is in a closed state. At this time, the microparticle sample pool (7) and the deionized water storage pool (8) take microparticle samples and deionized water from the microparticle sample inlet (10) and the second deionized water inlet (15) to be sucked into the depressible chamber (13). The pressing state of the previous step is repeated to achieve the automatic sample injection operation. (3) When the microparticle sample flows into the separation zone (16), it first flows into the secondary separation channel (20), and the signal generator (4) sets electrical parameters so that one particle in the mixed microparticle sample is adsorbed on the dielectrophoresis electrode (12) by the positive dielectrophoresis force, thereby achieving separation of one sample in the mixed microparticle sample; then, the remaining microparticle sample flows into the main separation channel (21), and is squeezed on both sides of the channel near the dielectrophoresis electrode (12) by the deionized water flowing in from the second deionized water inlet (15). At this time, the microparticles near the dielectrophoresis electrode (12) are separated into at least two other particles under the action of the negative dielectrophoresis force according to the difference in their own dielectric constants and the differences in particle size and morphology, and the separated particles flow out from the outlets (22) and flow into the particle collection chamber (6); (4) stopping the injection, pumping deionized water from the first deionized water inlet (11) to clean the separation area (16) and the secondary separation channel (20), and stopping the signal generator after the deionized water flows into the main separation channel (21), and the first particles adsorbed on the dielectrophoresis electrode (12) are detached, flow out from the outlet (22), and flow into the particle collection chamber (6); (5) Repeat steps (1) to (4) to achieve real-time high-throughput portable separation of multiple biological microparticles.

Images