CN107746794B - Cell separation device - Google Patents
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- CN107746794B CN107746794B CN201710894049.XA CN201710894049A CN107746794B CN 107746794 B CN107746794 B CN 107746794B CN 201710894049 A CN201710894049 A CN 201710894049A CN 107746794 B CN107746794 B CN 107746794B
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- B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
- B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
- B01L3/502761—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules
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- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
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- B01L2400/00—Moving or stopping fluids
- B01L2400/04—Moving fluids with specific forces or mechanical means
- B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
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- B01L2400/04—Moving fluids with specific forces or mechanical means
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- B01L2400/0409—Moving fluids with specific forces or mechanical means specific forces centrifugal forces
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- C12N2509/00—Methods for the dissociation of cells, e.g. specific use of enzymes
Abstract
The invention discloses a cell separation device which comprises a multi-stage microfluidic chip, wherein the multi-stage microfluidic chip comprises a first channel, a second channel and a third channel, the first channel and the second channel form a first passage, the first channel and the third channel form a second passage, one end of the first channel is provided with a first input port, one end of the second channel is provided with a second input port, the other end of the second channel is provided with a first output port and a second output port, one end of the third channel is provided with a third input port, and the other end of the third channel is provided with a third output port and a fourth output port. The invention also provides a method for separating cells by using the cell separation device. The invention adopts the multi-stage microfluidic chip in the blood cell separation device, can realize the simultaneous separation of four cells, has high flux and high speed, does not influence the activity of the separated cells, and has low cost, simple and convenient operation and easy popularization.
Description
Technical Field
The invention relates to the field of medical instruments, in particular to a cell separation device.
Background
Cancer-related mortality rates have steadily increased over the past decades and remain a major cause of human death. A study conducted by the World Health Organization (WHO) found that if cancer patients were diagnosed and treated before metastatic cancer occurred, a mortality rate of at least 30% was preventable. When Circulating Tumor Cells (CTCs) detach from a primary or metastatic tumor and flow into the peripheral blood stream, the tumor undergoes metastasis. CTCs were first established by the australian Thomas Ashworth in 1869. Because the CTCs in the peripheral blood of cancer patients have low occurrence frequency and high difficulty in accurate counting, the CTCs are not widely used for diagnosis and treatment. The potential role of CTCs in the process of metastasis is not fully understood. Therefore, effective and accurate methods for counting and characterizing CTCs are urgently needed to facilitate cancer diagnosis, prognosis and treatment.
During the last two decades, many researchers have explored efficient and reliable systems for the isolation of CTCs. Biological techniques based on cell biological characteristics such as specific expression of biomarkers, or physical properties such as size and deformability of CTCs have been developed to separate CTCs from whole blood in high yield and high purity. One of the most commonly used biological methods is the antibody-based technique (including the FDA-approved CellSearch system, Veridex).
The CellSearch system uses anti-EpCAM (epithelial cell adhesion molecule, specific for human breast cancer cells) conjugated magnetic beads for immunomagnetic capture and isolation of CTCs. This system is considered the gold standard for the isolation and enumeration of CTCs. However, given the presence of epithelial-mesenchymal transition (EMT) and the variation in EpCAM expression levels on different types of tumor cells, a proportion of CTCs may be lost during the isolation process. Therefore, the separation and counting of CTCs cannot be achieved with high accuracy. Furthermore, for subsequent studies on the collected circulating tumor cells, techniques that collect intact cells without labeling are more attractive.
Another class of methods commonly used are label-free isolated CTCs, including microfluidic filters, inertial focusing, acoustics, optics, and Dielectrophoresis (DEP). These methods take advantage of the larger and stiffer nature of CTCs than normal blood cells. However, these methods also have their limitations. Acoustic, optical and Dielectrophoresis (DEP) require additional working fields and longer processing times, whereas current inertial focusing methods can only separate one CTC from normal blood cells and are highly specific, with only specific devices capable of separating cells of a specific size.
Disclosure of Invention
The invention provides a cell separation device, which utilizes the property of fluid under the micron scale, namely, the fluid forms laminar flow under the micron scale, particles with different sizes occupy different balance positions in the laminar flow due to the balance of inertia lift force and Dean drag force, and cells can be separated simply, high-flux and high-efficiency.
In order to achieve the object of the present invention, one aspect of the present invention provides a cell separation device and a method of separating cells. In one embodiment, the cell separation device is characterized by comprising a multi-stage microfluidic chip, wherein the multi-stage microfluidic chip comprises a first channel, a second channel and a third channel, one end of the first channel is provided with a first input port, one end of the second channel is provided with a second input port, the other end of the second channel is provided with a first output port and a second output port, one end of the third channel is provided with a third input port, and the other end of the third channel is provided with a third output port and a fourth output port.
Furthermore, the first channel and the second channel of the multistage microfluidic chip are spiral single-conduit channels, the front half part of the third channel of the multistage microfluidic chip is a Z-shaped single-conduit channel, the rear half part of the third channel of the multistage microfluidic chip is a semicircular single-conduit channel, and the front half part of the third channel is a part connected to the third injection pump.
Further, the first channel further comprises a first diverging opening; the first input port is positioned at the innermost layer of the first channel;
the first branch opening is positioned at the outermost layer of the first channel, the first channel is respectively connected with the second channel and the third channel at the first branch opening through pipelines, the pipeline connected with the second channel is positioned at one side far away from the center of the first channel, and the pipeline connected with the third channel is positioned at one side close to the center of the first channel.
Further, the second channel also comprises a second bifurcation port and a third bifurcation port; the second branch port is positioned at the outermost layer of the second channel, and the second channel is connected with the second input port and the first channel through pipelines at the second branch port; the third bifurcation port is located at the innermost layer of the second channel and is provided with a first output port and a second output port, the first output port is located on one side far away from the center of the second channel, and the second output port is located on one side close to the center of the second channel.
Further, the third channel further comprises a fourth fork and a fifth fork; the third channel is connected with a third input port and the first channel through pipelines at the fourth branch port; the fifth fork opening is positioned at one end of the semicircular single conduit channel of the third channel, the third channel is provided with a third output port and a fourth output port at the fifth fork opening, the third output port is positioned at one side far away from the circle center of the semicircular single conduit channel, and the fourth output port is positioned at one side close to the circle center of the semicircular single conduit channel.
Further, the cross sections of the first channel, the second channel and the third channel are rectangular.
Furthermore, the first channel, the second channel and the third channel of the multistage microfluidic chip are positioned on the same horizontal plane.
Further, the cell separation device also comprises an injection pump controller, a first injection pump, a second injection pump and a third injection pump; the first input port is connected with the first injection pump through a plastic conduit, the second input port is connected with the second injection pump through a plastic conduit, the third input port is connected with the third injection pump through a plastic conduit, and the injection pump controller is respectively connected with the first injection pump, the second injection pump and the third injection pump through plastic conduits; the liquid injected by the first injection pump is a sample to be separated, and the liquid injected by the second injection pump and the third injection pump is buffer liquid.
Further, the flow rates of the first syringe pump, the second syringe pump, and the third syringe pump are controlled by the syringe pump controller.
In another aspect of the present invention, there is provided a novel method for separating cells, capable of simultaneously separating four kinds of cells using the cell separation device as described above, wherein a sample first enters a first channel and forms a laminar flow, and the cells are divided into a first cell part and a second cell part according to the size of the diameter by a balance of inertial lift force and Dean drag force; the first cell portion comprises two cells with smaller volume, the second cell portion comprises two cells with larger volume, and at the first fork opening, the first cell portion enters the second channel, and the second cell portion enters the third channel; separating two cells with different diameters in the first cell part through inertial lifting force and Dean dragging force in a second channel, and outputting the two cells through a first output port and a second output port; in the third channel, the second cell part is stressed in balance in the Z-shaped single guide pipe channel and is focused in the middle of the channel, two kinds of cells with different diameters in the second cell part are separated in the semicircular single guide pipe channel through centrifugal force and Dean drag force, and the two kinds of cells are output through a third output port and a fourth output port.
The cell separation device of the invention combines microfluidic channels with different shapes on a microfluidic chip for the first time to form a multi-stage inertial focusing microfluidic chip, and related documents and patents are not published before. By using the multi-stage inertial focusing microfluidic chip, cells with different sizes can be simultaneously and continuously separated by only utilizing the mechanical characteristics of fluid, and the separation efficiency and the precision are high.
When the cell separation device is used for separating cells, a sample only needs to be subjected to conventional centrifugation, supernatant is removed, and separation can be performed after the sample is resuspended by using a buffer solution without modifying the cells (such as endowing immunomagnetic beads), only the mechanical properties of fluid are utilized in the separation process, and external working fields (such as electric fields and magnetic fields) are not required, so that the separation method is simplified, the activity of the cells obtained by the separation of the multi-stage microfluidic chip is not influenced, complete cells can be obtained, and the separated cells can be used for further research. Wherein, through the velocity of flow of accurate control syringe pump of syringe pump controller, can realize the separation of the blood cell of equidimension not in a flexible way.
In addition, the multi-stage inertial focusing microfluidic chip has low cost, and the cost for separating cells is reduced.
The conception, the specific steps, and the technical effects produced by the present invention will be further described in conjunction with the accompanying drawings to fully understand the objects, the features, and the effects of the present invention.
Drawings
Fig. 1 is a schematic view of a blood cell separation apparatus according to an embodiment of the present invention.
Fig. 2 is a schematic view of a channel structure of a multi-stage microfluidic chip according to an embodiment of the present invention.
Fig. 3 is a schematic plan view of a channel of a multi-stage microfluidic chip according to an embodiment of the present invention.
FIG. 4 is a graph showing the force analysis of blood cells in the third channel according to the embodiment of the present invention.
FIG. 5 is a flow cytogram of output cells from each output port provided by an embodiment of the present invention.
The figures are labeled as follows:
1, a multistage microfluidic chip, 2, a syringe pump controller, 3, a first syringe pump, 4, a second syringe pump and 5, a third syringe pump;
11 a first channel, 12 a second channel, 13 a third channel;
111 first input port, 112 first split port, 121 second input port, 122 second split port, 123 third split port, 124 first output port, 125 second output port, 131 third input port, 132 fourth split port, 133 fifth split port, 134 third output port, 135 fourth output port.
Detailed Description
EXAMPLE 1 cell separation apparatus
A cell separation device, as shown in FIG. 1, comprises a multi-stage microfluidic chip 1, a syringe pump controller 2, a first syringe pump 3, a second syringe pump 4, and a third syringe pump 5.
As shown in fig. 1, the syringe pump controller 2 is connected to a first syringe pump 3, a second syringe pump 4, and a third syringe pump 5 through plastic tubes having an outer diameter of 1mm and an inner diameter of 0.8mm, respectively.
As shown in fig. 2 and 3, the multi-stage microfluidic chip 1 includes a first channel 11, a second channel 12, and a third channel 13, where the first channel 11 and the second channel 12 form a first channel, and the first channel 11 and the third channel 13 form a second channel. The first channel 11 and the second channel 12 are spiral single-conduit channels, the front half part of a third channel 13 of the multi-stage microfluidic chip 1 is a Z-shaped single-conduit channel, and the rear half part of the third channel is a semicircular single-conduit channel.
As shown in fig. 1 and 2, the first passage 11 further includes a first diverging port 112; the first input port 111 is located at the innermost layer of the first channel 11, the first input port 111 is connected with the first injection pump 3 through a plastic conduit, the outer diameter of the plastic conduit is 1mm, and the inner diameter of the plastic conduit is 0.8 mm; the first fork 112 is located at the outermost layer of the first channel 11, the first fork 112 divides the outermost end of the first channel 11 into two paths, and the two paths are respectively connected with the second channel 12 and the third channel 13 through pipelines, the pipeline connected with the second channel 12 is located at one side far away from the center of the first channel 11, and the pipeline connected with the third channel 13 is located at one side close to the center of the first channel 11.
As shown in fig. 1 and 2, the second channel 12 further includes a second bifurcation 122, a third bifurcation 123; the second branch port 122 is located at the outermost layer of the second channel 12, the second branch port 122 divides the tail end of the outermost layer of the second channel 12 into two paths, one path is connected with the second input port 121 through a pipeline, and the other path is connected with the first channel 11 through a pipeline; the second inlet 121 is connected to the second syringe pump 4 by a plastic conduit having an outer diameter of 1mm and an inner diameter of 0.8 mm; the third branch port 123 is located at the innermost layer of the second channel 12, the third branch port 123 divides the end of the innermost layer of the second channel 12 into two paths, one path has a first output port 124, the other path has a second output port 125, the path having the first output port 124 is located at one side far away from the center of the second channel 12, and the path having the second output port 125 is located at one side near the center of the second channel 12.
As shown in fig. 1 and 2, the third channel 13 further includes a fourth diverging opening 132 and a fifth diverging opening 133; the fourth branch port 132 is positioned at the tail end of the Z-shaped single-conduit passage of the third passage 13, the fourth branch port 132 divides the tail end of the Z-shaped single-conduit passage of the third passage 13 into two paths, one path is connected with the third input port 131 through a pipeline, and the other path is connected with the first passage 11 through a pipeline; the third inlet 131 is connected to the third syringe pump 5 via a plastic conduit having an outer diameter of 1mm and an inner diameter of 0.8 mm; the fifth fork 133 is located at the tail end of the semicircular single-conduit channel of the third channel 13, the fifth fork 133 divides the tail end of the semicircular single-conduit channel of the third channel 13 into two paths, one path is provided with a third output port 134, the other path is provided with a fourth output port 135, the path provided with the third output port 134 is located on one side far away from the circle center of the semicircular single-conduit channel, and the path provided with the fourth output port 135 is located on one side close to the circle center of the semicircular single-conduit channel.
The cross sections of the first channel 11, the second channel 12 and the third channel 13 are rectangular.
The first channel 11, the second channel 12 and the third channel 13 of the multi-stage microfluidic chip 1 are positioned on the same horizontal plane.
The cross section of the first channel 11 is rectangular, and the size is 300 multiplied by 80 μm; the second channel 12 has a rectangular cross section with dimensions of 600 x 80 μm; the cross-section of the third channel 13 is rectangular with dimensions of 200X 80 μm and the radius of the semicircular single-conduit channel is 6 mm.
The liquid injected by the first injection pump 3 is a sample to be separated, and the liquid injected by the second injection pump 4 and the third injection pump 5 is a buffer solution. The flow rates of first syringe pump 3, second syringe pump 4, and third syringe pump 5 are controlled by syringe pump controller 2.
Example 2 method for isolating cells
In this example, it is necessary to separate human breast cancer cells (MCF-7), human lung cancer cells (A549), erythrocytes and leukocytes in blood.
In other embodiments, other cells may also be isolated by adjusting the flow rate of the syringe pump or the size of the chip.
1. Preparation of blood samples
Centrifuging 400g of 1mL of blood for 20min at 20 ℃ and reducing the speed to 5g/s, taking 200 mu L of neutrophils and red blood cells, resuspending in 800 mu L of cell frozen stock solution, and storing in a refrigerator at-80 ℃ for later use.
2. Blood cell separation
Resuscitated white blood cells and red blood cells and human lung cancer cells (A549) and human breast cancer cells (MCF-7) are treated according to the proportion of 5 x 105/mL，5×105/mL，2×104/mL，and4×104The cell suspension (4 mL) was prepared by suspending in physiological saline solution, and was injected into the first input port 111 through the first syringe pump 3, 10mL of 0.09% physiological saline was injected into the second input port 121 through the second syringe pump 4, and 10mL of 0.09% physiological saline was injected into the third input port 131 through the third syringe pump 5. The flow rate of the first syringe pump 3 was set to 2mL/min, and the flow rates of the second syringe pump 4 and the third syringe pump 5 were set to 1.2 mL/min. Four cells in blood: after red blood cells, white blood cells, human breast cancer cells (MCF-7) and human lung cancer cells (A549) are separated by the first channel 11, the white blood cells and the red blood cells enter a pipeline connected with the second channel 12 at the first fork 112, and the human breast cancer cells (MCF-7) and the human lung cancer cells (A549) enter a pipeline connected with the third channel 13. After separation through the second channel 12, in a third bifurcationAnd the port 123, the red blood cells enter the first output channel 126 and are output through the first output port 124, and the white blood cells enter the second output channel 127 and are output through the second output port 125. After being separated through the third channel 13, human breast cancer cells (MCF-7) are output through the fourth output port 135 and human lung cancer cells (a549) are output through the third output port 134 at the fifth divergence 133. 1mL of blood can be separated within 30min, and the output cells from each output port are collected and the flow cytogram is prepared for cell separation as shown in FIG. 5. FIG. 5, panel A, is a control, flow cytogram of unseparated cells, with total red 35.16%, white 50.23%, 5.57% and 8.02% human lung cancer cells (A549), and breast cancer cells (MCF-7); in fig. 5B is a flow cytogram of the first output port output cells, wherein the red blood cells account for 94.78% of the population; fig. 5C is a flow cytogram of the second output port output cells, wherein red blood cells account for 15.92% of the population and white blood cells account for 80.79% of the population; in fig. 5D is a flow cytogram of third output port output cells, wherein human lung cancer cells (a549) account for 75.04% of the population; FIG. 5E is a flow cytogram of fourth output port output cells, in which human breast cancer cells (MCF-7) account for 84.4% of the total; in fig. 5, F is a bar chart prepared according to the total percentage of four cells in the unseparated cells and the output cells of each output port, and the cells corresponding to the four bars from left to right in each group are erythrocytes, leukocytes, human lung cancer cells (a549) and human breast cancer cells (MCF-7), and the cell separation rate is greater than 75% according to the above results.
3 stress analysis of blood cells in multi-stage microfluidic chip
In the invention, the property of the fluid under the micron scale is utilized to realize the continuous separation of four cells with different sizes. At the micrometer scale, the fluid forms laminar flow, and particles with different sizes occupy different balance positions in the laminar flow due to the balance of inertial lift force and Dean drag force. The cells in the two spiral channels of the first channel 11 and the second channel 12 are subjected to inertial lift and Dean drag. These two forces are related to the size of the diameter of the cell, the velocity of the fluid and the cross-sectional area of the channel. The equilibrium position of the cell is related to the size of the cell under certain velocity and cross section. Cells with a large diameter occupy equilibrium positions close to the inside of the channel, and cells with a small diameter occupy equilibrium positions far from the inside.
The diameter sizes of the four separated cells are as follows from large to small: human breast cancer cells (MCF-7), human lung cancer cells (A549), white blood cells and red blood cells. Within the first channel 11, human breast cancer cells (MCF-7) and human lung cancer cells (a549) occupy equilibrium positions near the inside of the channel, while red blood cells and white blood cells occupy equilibrium positions far from the inside of the channel. So that at the first divergence orifice 112, human breast cancer cells (MCF-7) and human lung cancer cells (a549) flow into the third channel 13 and red blood cells and white blood cells flow into the second channel 12. In the second channel 12, the flow rate is changed by the buffer liquid injected by the second syringe pump 4, so that the white blood cells and the red blood cells occupy respective equilibrium positions, the white blood cells occupy an equilibrium position close to the inner side of the channel, and the red blood cells occupy a position far from the inner side of the channel, so that, at the third branch port 123, the red blood cells are output from the first output port 124 after entering the first output channel, and the white blood cells are output from the second output port 125 after entering the second output channel.
In the third channel 13, the flow rate is changed by the buffer solution injected by the third syringe pump 5. As shown in fig. 4, human breast cancer cells (MCF-7) and human lung cancer cells (a549) were force-balanced in the Z-channel, focusing in the middle of the channel. Flows into the semicircular channel where the equilibrium position of the cells is mainly determined by the centrifugal force and Dean drag. When the flow rate is low, the cells maintain their original equilibrium position, flowing along the middle of the channel. With a slow increase in flow rate, centrifugal force dominates, and human lung cancer cells (a549) begin to approach the inside of the channel, human breast cancer cells (MCF-7) maintain an equilibrium position at an intermediate position. When the flow rate is higher, Dean drag dominates, human breast cancer cells (MCF-7) begin to approach the medial side, while human lung cancer cells (a549) begin to flow to the lateral side, maintaining their respective equilibrium positions until fifth bifurcation 133. When the flow rate of the third syringe pump 5 is set to 1.2mL/min, the separated human breast cancer cells (MCF-7) are output from the fourth output port 135, and the human lung cancer cells (a549) are output from the third output port 134.
In this embodiment, three different-shaped inertia-based microfluidic channels are integrated into one chip, enabling size-based, label-free, high-throughput, and efficient separation of CTCs in blood cells. The device shown in this example integrates multiple channels onto one chip, and separates four different types of cells simultaneously, namely human breast cancer cells (MCF-7), human lung cancer cells (A549), erythrocytes and leukocytes. Research results show that the design simplifies a method for separating cells, has higher separation efficiency, can simultaneously separate and collect four types of intact cells, and the separated cells can be continuously used for subsequent research.
The foregoing detailed description of the preferred embodiments of the invention has been presented. It should be understood that numerous modifications and variations could be devised by those skilled in the art in light of the present teachings without departing from the inventive concepts. Therefore, the technical solutions available to those skilled in the art through logic analysis, reasoning and limited experiments based on the prior art according to the concept of the present invention should be within the scope of protection defined by the claims.
Claims (6)
1. A cell separation device is characterized by comprising a multi-stage microfluidic chip, wherein the multi-stage microfluidic chip comprises a first channel, a second channel and a third channel, the first channel and the second channel are spiral single-conduit channels, the front half part of the third channel is a Z-shaped single-conduit channel, and the rear half part of the third channel is a semicircular single-conduit channel; the first channel and the second channel form a first passage, the first channel and the third channel form a second passage, one end of the first channel is provided with a first input port, one end of the second channel is provided with a second input port, the other end of the second channel is provided with a first output port and a second output port, one end of the third channel is provided with a third input port, the other end of the third channel is provided with a third output port and a fourth output port, and the first channel further comprises a first branch port; the first input port is located at the innermost layer of the first channel; the first branch port is positioned at the outermost layer of the first channel, the first channel is respectively connected with the second channel and the third channel at the first branch port through pipelines, the pipeline connected with the second channel is positioned at one side far away from the center of the first channel, and the pipeline connected with the third channel is positioned at one side close to the center of the first channel; the second channel further comprises a second bifurcation port and a third bifurcation port; the second branch port is positioned at the outermost layer of the second channel, and the second channel is respectively connected with the second input port and the first channel at the second branch port through pipelines; the third bifurcation is positioned at the innermost layer of the second channel and is provided with a first output port and a second output port, the first output port is positioned at one side far away from the center of the second channel, and the second output port is positioned at one side close to the center of the second channel; one end of the Z-shaped single conduit of the third channel is provided with a fourth fork opening, and the fourth fork opening is respectively connected with a third output port and the first channel through conduits; one end of the semicircular single-conduit passage of the third channel is provided with a fifth fork, the fifth fork is provided with a third output port and a fourth output port, the third output port is positioned on one side far away from the circle center of the semicircular single-conduit passage, and the fourth output port is positioned on one side close to the circle center of the semicircular single-conduit passage.
2. The cell separation device of claim 1, wherein the first channel, the second channel, and the third channel are rectangular in cross-section.
3. The cell separation device according to claim 1, wherein the first channel, the second channel, and the third channel of the multi-stage microfluidic chip are located at the same horizontal plane.
4. The cell separation device of claim 1, further comprising a syringe pump controller, a first syringe pump, a second syringe pump, a third syringe pump; the first input port is connected with a first injection pump through a plastic conduit, the second input port is connected with a second injection pump through a plastic conduit, the third input port is connected with a third injection pump through a plastic conduit, and the injection pump controller is respectively connected with the first injection pump, the second injection pump and the third injection pump through plastic conduits; the liquid injected by the first injection pump is a sample to be separated, and the liquid injected by the second injection pump and the liquid injected by the third injection pump are buffer solutions.
5. The cell separation apparatus of claim 4, wherein flow rates of the first syringe pump, the second syringe pump, and the third syringe pump are controlled by the syringe pump controller.
6. A method for separating cells, which is capable of simultaneously separating four kinds of cells using the cell separation device according to any one of claims 1 to 5, wherein a sample first enters the first channel and forms a laminar flow, and the cells are divided into a first cell fraction and a second cell fraction according to the size of the diameter by a balance of inertial lift force and Dean drag force; the first cell fraction comprises two cells of a smaller volume and the second cell fraction comprises two cells of a larger volume, the first cell fraction entering the second channel at the first bifurcation and the second cell fraction entering the third channel; separating two cells with different diameters in the first cell part through inertial lift force and Dean drag force in the second channel, and outputting the two cells through the first output port and the second output port; in the third channel, the second cell part is stressed in a Z-shaped single guide pipe channel in a balanced manner, is focused in the middle of the channel, and in a semicircular single guide pipe channel, two kinds of cells with different diameters in the second cell part are separated through centrifugal force and Dean drag force, and the two kinds of cells are output through the third output port and the fourth output port.
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| CN115637257B (en) * | 2022-12-07 | 2023-04-14 | 翔鹏佑康(北京)科技有限公司 | Circulating tumor cell screening method based on inertial focusing microfluidics |
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