CN111172031A - Cell capturing and screening method - Google Patents

Cell capturing and screening method Download PDF

Info

Publication number
CN111172031A
CN111172031A CN202010055458.2A CN202010055458A CN111172031A CN 111172031 A CN111172031 A CN 111172031A CN 202010055458 A CN202010055458 A CN 202010055458A CN 111172031 A CN111172031 A CN 111172031A
Authority
CN
China
Prior art keywords
flow channel
cell
capturing
screening
chip
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
CN202010055458.2A
Other languages
Chinese (zh)
Inventor
许行尚
杰弗瑞·陈
孙威严
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Nanjing Lanyu Biological Technology Co Ltd
Original Assignee
Nanjing Lanyu Biological Technology Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Nanjing Lanyu Biological Technology Co Ltd filed Critical Nanjing Lanyu Biological Technology Co Ltd
Priority to CN202010055458.2A priority Critical patent/CN111172031A/en
Publication of CN111172031A publication Critical patent/CN111172031A/en
Pending legal-status Critical Current

Links

Images

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/16—Microfluidic devices; Capillary tubes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/58—Reaction vessels connected in series or in parallel
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
    • C12M33/04—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus by injection or suction, e.g. using pipettes, syringes, needles
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
    • C12M47/04—Cell isolation or sorting

Abstract

The invention discloses a method for capturing and screening cells, which specifically comprises the following steps: the cell capturing and screening method comprises the following steps of (1) adopting at least one cell capturing and screening chip, adding cell suspension to be screened from a sample inlet of the cell capturing and screening chip, enabling the cell suspension to enter a flow channel of the cell capturing and screening chip, (2) enabling the cell suspension to flow to a sample outlet of the cell capturing and screening chip in the flow channel until the cell suspension after reaction flows to the sample outlet of the cell capturing and screening chip, and (3) enabling the cell suspension to flow out of a screened cell or cell compound from the sample outlet.

Description

Cell capturing and screening method
Technical Field
The invention belongs to the technical field of cell biology experimental devices, and particularly relates to a cell capturing and screening method and a cell capturing and screening chip adopted by the same.
Background
In recent years, new antibody drugs have shown good therapeutic effects in the treatment of serious diseases such as malignant tumors and autoimmune diseases, and the development of antibody drugs in China and abroad has become a new focus and is extremely competitive. In the development process of new antibody drugs, a plurality of key technologies need to be established and applied, including antibody screening and function confirmation, cell strain construction and process development, pilot plant process amplification, preclinical research and the like. At present, in the aspect of antibody screening and process development in China, a plurality of technical difficulties exist, for example, most of the currently adopted antibody screening methods by a hybridoma cell technology have a plurality of defects, large-throughput screening cannot be realized, sequences and affinity data of tens of thousands of antibodies cannot be simultaneously obtained, and the probability of obtaining an optimized antibody sequence is low. Therefore, it is necessary to establish a cell capturing and screening method that can realize large-throughput screening and more effectively realize refinement.
Cells are the basic unit of life body structure and life activity, and are the basis of life science and biomedical research. Cell screening provides fundamental information for studies of drug screening based on cell analysis, intracellular gene expression, and the like. The rapid, high-throughput and label-free cell screening method not only can provide great convenience for biological research, but also can provide direct help for biomedical research, and is beneficial to the research of life processes and the early diagnosis of serious diseases.
The current cell screening methods mainly comprise a laser method, a fluorescence labeling method and a label-free method based on optical trapping force.
The laser focusing beam array process of the laser method is relatively slow, the method deflects one type of cells by a certain angle and then enters a single collection storage pool, and other types of cells directly enter another storage pool, so that the cell screening is realized, but the method has the defects that when the cell types are different, the cells can be continuously deflected in space, so that the separation of the cells is not obvious; therefore, this method is not suitable for high-throughput, high-sensitivity cell screening; the fluorescence labeling method has obvious limitation and may influence the activity of cells; on the other hand, the label-free method based on the optical trapping force has problems of low flux, low sensitivity, and the like.
Chinese patent document (application number: 201410131357.3) discloses a microfluidic chip for cell screening. This chip includes T type groove, main channel part and branch passageway, and T type groove includes inlet channel and 2 sheath flow channels, its characterized in that two subchannels are separated out to the end of main channel: the tail end of one sub-channel forms four guide channels which are connected in a ray shape, and the other sub-channel is connected with a waste liquid pool; each of the four guide channels is spaced at 30 degrees; the branch channel connected with each sub-channel comprises two 90-degree bent angles which are distributed anticlockwise, the initial part of the branch channel connected with the sub-channel is perpendicular to the main channel, then the anticlockwise 90-degree turning is performed twice in sequence to form the branch channel, the tail end of the branch channel is connected with 2 cell collectors, the No. 2 cell collector is connected with the tail end of the branch channel in parallel, and the No. 1 cell collector is connected with the tail end of the branch channel at an angle of 30 degrees. The disadvantage of this invention is that high throughput and high sensitivity cell screening using laser methods is still not achieved.
Chinese patent document (application No. 201610398852.X) discloses a microfluidic device and method for size-based detection of circulating tumor cells, the device comprising: the device comprises a solution storage chamber, a micro-fluidic chip, a waste liquid collecting needle cylinder and a power system which are connected in sequence; the micro-fluidic chip is formed by laminating a glass substrate layer and a PDMS chip layer, wherein the PDMS chip layer comprises sample inlets which are sequentially communicated; a cake filtration zone formed by an array of pillars; the target cell screening area comprises a plurality of main pipelines and side pipelines which extend in parallel at intervals and are communicated through a filtering channel, the front end of each main pipeline is open, the rear end of each main pipeline is provided with the filtering channel, the front end of each side pipeline is closed, the rear end of each side pipeline is open, the main pipelines and the side pipelines have first heights larger than the size of the circulating tumor cells, and the filtering channels have second heights smaller than the size of the circulating tumor cells; and a sample outlet. The invention has the disadvantages of complicated chip structure, more and compact flow channel distribution, complicated manufacturing process, high cost, no contribution to batch production, no realization of free combination of multiple chips and limited cell screening types.
Therefore, there is a need to develop a method for capturing and screening cells, which has high sensitivity, high throughput, simple chip structure, convenient operation, low cost, and flexible combination of chip.
Disclosure of Invention
The invention aims to solve the technical problem of providing a cell capturing and screening method which has the advantages of high sensitivity, large flux, simple chip structure, convenient operation and low cost and can flexibly combine and use the chip.
In order to solve the technical problems, the invention adopts the technical scheme that the method for capturing and screening the cells specifically comprises the following steps:
(1) adding a cell suspension to be screened from a sample inlet of the cell capturing and screening chip by adopting at least one cell capturing and screening chip, wherein the cell suspension enters a flow channel of the cell capturing and screening chip;
(2) the cell suspension flows to the sample outlet of the cell capturing and screening chip in the flow channel until the reacted cell suspension flows to the sample outlet of the cell capturing and screening chip;
(3) and (4) flowing out the screening cells or cell complexes from the sample outlet.
By adopting the technical scheme, the cell suspension to be screened is added from the sample inlet, after entering the flow channel, the cell suspension flows forwards automatically inside or under the control of the pump, and can react with the substances fixed on the wall of the flow channel or in the cavity, the cell suspension after reaction flows forwards continuously through the flow channel, and can continuously react with the substances on the wall of the subsequent flow channel or in the cavity in sequence, finally the cell or cell compound is screened by flowing out of the sample outlet, thereby realizing the screening of the cells, and solving the problems of low flux, low sensitivity and the like, wherein the cell capturing and screening chip can be different in structure, area, volume and flow speed, and can also be combined by a plurality of cell capturing and screening chips with different structures in series for use, the substances are pre-fixed in the wall of the flow channel or the cavity, and can react with and combine with impurity cells to adsorb the impurity cells and the like, collecting and detecting the cell to be screened after flowing out; or the pre-fixed substance reacts and combines with the cells to be screened, and after impurity cells and the like flow out, the cells to be screened are eluted and collected for detection; ensure that the cells to be screened flow out from the sample outlet through the flow channel, thereby increasing the sensitivity of cell screening.
As a preferred embodiment of the present invention, in the step (2), at least one pre-immobilized substance in the flow channel wall or/and the cavity is reacted during the flow process, until the reacted cell suspension flows to the sample outlet of the cell capture and screening chip. A cavity connected with the flow channel can be arranged in the cell capturing and screening chip, the wall of the flow channel is pre-coated with an adsorption substance to adsorb impurity cells, the cavity is internally provided with an adsorption substance or a fixed fluorescence labeling reaction substance, the fluorescence labeling substance reacts with the impurity cells or the cells to be screened, the cells to be screened or the impurity cells flow out after reacting, and the screening efficiency of the cells is detected; detecting the amount of the cells to be screened or reacting with impurity cells to indirectly reflect the amount of the cells to be screened, namely, the cell suspension after reaction continuously flows forwards through a flow channel, sequentially reacts in a subsequent flow channel and a cavity, and finally flows out the cells to be screened or cell compound from a sample outlet, or directly or indirectly detects the amount or screening rate of the cells to be screened through a fluorescent substance. The length of the flow channel of the cell capturing and screening chip is determined according to the requirement of screening rate.
As a preferred technical scheme of the invention, the method for capturing and screening the cells adopts at least two chips for capturing and screening the cells, and the adjacent two chips for capturing and screening the cells are connected in series. The cell capturing and screening chips can be used in series combination, and the length of the flow channel of the cell capturing and screening chips and the number of the series connection of the cell capturing and screening chips are determined according to the requirement of the screening rate.
As the preferred technical scheme of the invention, in the step (1), the cell suspension to be screened is controlled by a matched detection instrument or added into the sample inlet by adopting a liquid transfer device.
The invention is further improved in that the cell capturing and screening chip comprises a sample inlet, a sample outlet and a flow channel, wherein the sample inlet is communicated with the sample outlet through the flow channel; the flow channel is provided with a plurality of corners from the sample inlet to the sample outlet. The cell capturing and screening chip is formed by laminating a glass substrate and a polydimethylsiloxane PDMS chip layer, wherein the height and the width of a flow channel are determined according to the requirement of screening rate.
As a preferred technical scheme of the invention, the flow channel between two adjacent corners is linear. The flow channel is set to be a linear type, so that the optimization of cell flow and the length of the flow channel can be realized to the greatest extent, and the cell can be screened by utilizing the particle size of the cell, namely, the flow channel is sequentially connected with chips, the width and the height of which are gradually reduced to allow the cell to be screened to pass through.
As a preferable technical scheme of the invention, the flow channels and the cavities are alternately connected in series, and the distances of the flow channels between the adjacent cavities at two non-corners are the same. Cavities are uniformly arranged among the linear flow channels, so that the cell capacity is increased, and meanwhile, adsorption substances can be arranged in the cavities, antibodies can also be fixed for immunoreaction, or other reaction substances are fixed, so that cells to be screened or impurity cells react to generate a new cell compound, and then, the cells to be screened or the new cell compound thereof are collected or detected in a later stage; the cell capturing and screening chip of the serial cavity flow channel can also be combined with the cell capturing and screening chips of other flow channels according to the screening requirement.
As a preferred technical solution of the present invention, the flow channel includes a first main flow channel, a second main flow channel, and a surface cavity flow channel, one end of the first main flow channel is connected to the sample inlet, the other end of the first main flow channel extends in a direction perpendicular to the first main flow channel toward the sample outlet and is connected to the surface cavity flow channel, the surface cavity flow channel is further connected to an extending end of the second main flow channel in a direction perpendicular to the second main flow channel toward the sample inlet, and the other end of the second main flow channel is connected to the sample outlet. The flow channel is arranged to be a surface cavity flow channel between the first main flow channel and the second main flow channel, so that the cavity can be completely filled with liquid without bubbles; meanwhile, the arrangement of the surface cavity flow channel increases the cell capacity, and is suitable for screening high-flux sample cell suspension; the cell capturing and screening chip of the surface cavity flow channel can also be combined with the cell capturing and screening chips of other flow channels according to the screening requirement.
As a preferred technical scheme, the width range of the flow channel is 20-500 mu m, the height range of the flow channel is 20-500 mu m, and the width-to-height ratio of the flow channel is 1: 1-2: 1; the cell sap carrying capacity of the flow channel is 1-2500 mu L.
As a preferred technical scheme of the invention, the diameter of the cavity is 1000-6500 mu m, and the ratio of the depth of the cavity to the diameter of the cavity is 1: 1-1: 2.
As a preferred technical scheme, the surface cavity flow channel is a hexagonal flow channel, upward-protruding columns are arranged inside the hexagonal flow channel, and the area of the hexagonal flow channel is 100-1000 mm2(ii) a The height of the hexagonal flow channel is 0.1-2 mm, and the cell liquid carrying capacity of the hexagonal flow channel is 10-2000 mu L. The effect of setting up the protruding post of upwards is to guarantee that the degree of depth of face cavity runner is unanimous, and the quantity of post is 4.
Compared with the prior art, the invention has the beneficial effects that:
(1) the cell capturing and screening chip with 3 structural flow channels is designed, one structure can be selected independently for use, and a plurality of structures can be connected in series for use;
(2) the cell capturing and screening chip can be controlled by a detection instrument or directly used by adding liquid by a liquid shifter;
(3) the cell capturing and screening method has the advantages of high cell screening sensitivity, large flux, simple structure, convenient operation and low cost, and can be flexibly combined for use;
(4) the method has wide application range, and can finally obtain the cells to be screened by two modes of adsorbing impurity cells and adsorbing the cells to be screened.
Drawings
The following further detailed description of embodiments of the invention is made with reference to the accompanying drawings:
FIG. 1 is a structural view of embodiment 4 of the present invention;
FIG. 2 is a structural view of embodiment 5 of the present invention;
FIG. 3 is a structural view of embodiment 6 of the present invention;
FIG. 4 is a structural view of embodiment 7 of the present invention;
FIG. 5 is a structural view of embodiment 8 of the present invention;
FIG. 6 is a structural view of embodiment 9 of the present invention;
wherein 1-a sample inlet; 2-a sample outlet; 3-a flow channel; 301-a primary channel one; 302-main runner two; 303-face cavity flow channel; 304-an extension end; 4-a cavity; 5-column.
Detailed Description
Example 1: the method for capturing and screening the cells specifically comprises the following steps:
(1) adopting a cell capturing and screening chip with a linear flow channel 3, and adding a cell suspension to be screened into the sample inlet 1 under the control of a matched detection instrument or by adopting a liquid transfer device, wherein the cell suspension enters the flow channel 3 of the cell capturing and screening chip;
(2) the cell suspension flows to the sample outlet 2 of the cell capturing and screening chip in the flow channel until the reacted cell suspension flows to the sample outlet 2 of the cell capturing and screening chip;
(3) and (3) flowing out the screened cells or cell complexes from the sample outlet 2.
The cell suspension to be screened is added from the sample inlet 1, the cell suspension flows forwards automatically inside or under the control of a pump after entering the flow channel 3 and can react with substances fixed on the wall of the flow channel, the cell suspension after reaction continuously flows forwards through the flow channel 3 and can continuously react with the substances on the wall of the subsequent flow channel in sequence, and finally the cell or cell compound is screened after flowing out of the sample outlet, so that the cell is screened, and the problems of low flux, low sensitivity and the like are solved.
Example 2: the method for capturing and screening the cells specifically comprises the following steps:
(1) adopting a cell capturing and screening chip with a flow channel 3 in a straight line and cavities alternately connected in series, and adding a cell suspension to be screened into the sample inlet 1 under the control of a matched detection instrument or by adopting a liquid transfer device, wherein the cell suspension enters the flow channel 3 of the cell capturing and screening chip;
(2) the cell suspension flows to a sample outlet 2 of the cell capturing and screening chip in the flow channel, and sequentially reacts with a plurality of pre-fixed substances on the wall of the flow channel and pre-fixed substances in a plurality of cavities 4 in the flow process, the reacted cell suspension continuously flows forwards through the flow channel 3, and sequentially reacts in the subsequent flow channel and cavities, and finally flows out of cells to be screened or cell compounds from the sample outlet, wherein the wall of the flow channel pre-wraps adsorbed substances, adsorbs impurity cells, a fluorescence labeling reaction substance is fixed in the cavity, the fluorescence labeling substance reacts with the impurity cells or the cells to be screened, the cells to be screened or the impurity cells react and then flow out, and the screening efficiency of the cells to be screened is detected; until the cell suspension after reaction flows to a sample outlet of the cell capturing and screening chip;
(3) and (4) flowing out the screening cells or cell complexes from the sample outlet.
Example 3: the method for capturing and screening the cells specifically comprises the following steps:
(1) two cell capturing and screening chips are connected in series, wherein a flow channel 3 of one cell capturing and screening chip is linear, a flow channel 3 of the other cell capturing and screening chip is linear and is alternately connected with a cavity in series, a cell suspension to be screened is added into the sample inlet by a matched detection instrument or a liquid transfer device, and the cell suspension enters the flow channel of the cell capturing and screening chip;
(2) the cell suspension flows to the sample outlet 2 of the cell capturing and screening chip in the flow channel 3, and sequentially reacts with a plurality of pre-fixed substances on the wall of the flow channel and pre-fixed substances in a plurality of cavities 4 in the flow process, the reacted cell suspension continuously flows forwards through the flow channel, sequentially reacts in the subsequent flow channel and cavities, and finally flows out of cells or cell compounds to be screened from the sample outlet, wherein the wall of the flow channel is pre-coated with adsorbed substances to adsorb impurity cells, fluorescent labeled reaction substances are fixed in the cavities 4, the fluorescent labeled substances react with the cells to be screened, the cells or the impurity cells react and then flow out, and the screening efficiency of the cells to be screened is detected; until the cell suspension after reaction flows to the sample outlet 2 of the cell capturing and screening chip;
(3) and (4) flowing out the screening cells or cell complexes from the sample outlet.
Example 4: as shown in fig. 1, the cell capturing and screening chip includes a sample inlet 1, a sample outlet 2 and a flow channel 3, wherein the sample inlet 1 is communicated with the sample outlet 2 via the flow channel 3; the flow channel 3 has 14 corners from the sample inlet 1 to the sample outlet 2; the cell capturing and screening chip is formed by laminating a glass substrate and a polydimethylsiloxane PDMS chip layer, and a flow channel 3 between two adjacent corners is linear; wherein the width range of the flow channel 3 is 200 μm, the height range of the flow channel 3 is 200 μm, and the width-to-height ratio of the flow channel 3 is 1: 1; the cell fluid loading of the flow channel 3 was 25. mu.L.
Example 5: as shown in fig. 2, the cell capturing and screening chip includes a sample inlet 1, a sample outlet 2 and a flow channel 3, wherein the sample inlet 1 is communicated with the sample outlet 2 via the flow channel 3; the flow channel 3 has 8 corners from the sample inlet 1 to the sample outlet 2; the cell capturing and screening chip is formed by laminating a glass substrate and a polydimethylsiloxane PDMS chip layer, and a flow channel 3 between two adjacent corners is linear; the flow channels 3 and the cavities 4 are alternately connected in series, and the distances of the flow channels between the adjacent cavities 4 at two non-corners are the same; wherein the width range of the flow channel 3 is 200 μm, the height range of the flow channel 3 is 200 μm, and the width-to-height ratio of the flow channel 3 is 1: 1; the cell liquid loading capacity of the flow channel 3 is 300 mu L; the diameter of the cavity 4 is 1000 microns, and the ratio of the depth of the cavity 4 to the diameter of the cavity 4 is 1: 1; wherein the cell carrying capacity is related to the width and height of the flow channel 3, the size of the cavity and the number of the cavities.
Example 6: as shown in fig. 3, the cell capturing and screening chip includes a sample inlet 1, a sample outlet 2 and a flow channel 3, wherein the sample inlet 1 is communicated with the sample outlet 2 via the flow channel 3; the flow channel 3 has 4 corners from the sample inlet 1 to the sample outlet 2; the cell capturing and screening chip is formed by attaching a glass substrate and a polydimethylsiloxane PDMS chip layer, the flow channel 3 comprises a first main flow channel 301, a second main flow channel 302 and a surface cavity flow channel 303, one end of the first main flow channel 301 is connected with the sample inlet 1, the other end of the first main flow channel 301 extends towards the direction of the sample outlet 2 along the direction vertical to the first main flow channel 301 and is connected with the surface cavity flow channel 303, and the surface cavity flow channel 303 is also connected with the second main flow channel 302 along the direction vertical to the second main flow channel 302The extending end 304 in the direction of the sample inlet 1 is connected, and the other end of the second main channel 302 is connected with the sample outlet 2; the surface cavity runner 303 is a hexagonal runner, the hexagonal runner is internally provided with a column 5 protruding upwards, and the area of the hexagonal runner is 500mm2(ii) a The height of the hexagonal flow channel is 0.5mm, and the cell liquid carrying capacity of the hexagonal flow channel is 250 mu L; the function of setting up upwards protruding post 5 is to guarantee that the degree of depth of face cavity runner is unanimous, and the quantity of post 5 is 4.
Example 7: as shown in fig. 4, the difference from the example 4 is that the aspect ratio of the flow channel 3 is 2: 1; the cell liquid loading capacity of the flow channel 3 is 75 mu L; specifically, the cell capturing and screening chip comprises a sample inlet 1, a sample outlet 2 and a flow channel 3, wherein the sample inlet 1 is communicated with the sample outlet 2 through the flow channel 3; the flow channel 3 has 14 corners from the sample inlet 1 to the sample outlet 2; the cell capturing and screening chip is formed by laminating a glass substrate and a polydimethylsiloxane PDMS chip layer, and a flow channel 3 between two adjacent corners is linear; wherein the width range of the flow channel 3 is 400 μm, the height range of the flow channel 3 is 200 μm, and the width-to-height ratio of the flow channel 3 is 2: 1; the cell fluid loading of the flow channel 3 was 75 μ L.
Example 8: as shown in fig. 5, the difference from the embodiment 5 is that the number and size of the cavities 4 are different, and the number of the cavities 4 is 13; the diameter of the cavity 4 is 3000 micrometers; specifically, the cell capturing and screening chip comprises a sample inlet 1, a sample outlet 2 and a flow channel 3, wherein the sample inlet 1 is communicated with the sample outlet 2 through the flow channel 3; the flow channel 3 has 4 corners from the sample inlet 1 to the sample outlet 2; the cell capturing and screening chip is formed by laminating a glass substrate and a polydimethylsiloxane PDMS chip layer, and a flow channel 3 between two adjacent corners is linear; the flow channels 3 and the 33 cavities 4 are alternately connected in series, and the distances of the flow channels between the adjacent cavities 4 at two non-corners are the same; wherein the width range of the flow channel 3 is 200 μm, the height range of the flow channel 3 is 200 μm, and the width-to-height ratio of the flow channel 3 is 1: 1; the cell liquid loading capacity of the flow channel 3 is 1000 mu L; the diameter of the cavity 4 is 3000 microns, and the ratio of the depth of the cavity 4 to the diameter of the cavity 4 is 1: 1.
Example 9: as shown in fig. 6, the difference from the embodiment 8 lies in that the number and size of the cavities 4 are different, and the number of the cavities is 10; the diameter of the cavity 4 is 6000 mu m; specifically, the cell capturing and screening chip comprises a sample inlet 1, a sample outlet 2 and a flow channel 3, wherein the sample inlet 1 is communicated with the sample outlet 2 through the flow channel 3; the flow channel 3 has 4 corners from the sample inlet 1 to the sample outlet 2; the cell capturing and screening chip is formed by laminating a glass substrate and a polydimethylsiloxane PDMS chip layer, and a flow channel 3 between two adjacent corners is linear; the flow channel 3 and the 33 cavities are alternately connected in series, and the distances of the flow channels between the adjacent cavities 4 at two non-corners are the same; wherein the width range of the flow channel 3 is 200 μm, the height range of the flow channel 3 is 200 μm, and the width-to-height ratio of the flow channel 3 is 1: 1; the cell liquid loading capacity of the flow channel 3 is 1000 mu L; the diameter of the cavity 4 is 6000 microns, and the ratio of the depth of the cavity 4 to the diameter of the cavity 4 is 1: 1.
The foregoing illustrates and describes the principles, general features, and advantages of the present invention. It will be understood by those skilled in the art that the present invention is not limited by the foregoing embodiments, and the foregoing embodiments and descriptions are only illustrative of the principles of the present invention, and that various changes and modifications, such as changes in the number of the cell capturing and screening chips and the shape and size of the flow channels of the cell capturing and screening chips, may be made without departing from the spirit and scope of the present invention, and such changes and modifications are within the scope of the present invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.

Claims (11)

1. A method for capturing and screening cells is characterized by comprising the following steps:
(1) adding a cell suspension to be screened from a sample inlet of the cell capturing and screening chip by adopting at least one cell capturing and screening chip, wherein the cell suspension enters a flow channel of the cell capturing and screening chip;
(2) the cell suspension flows to the sample outlet of the cell capturing and screening chip in the flow channel until the reacted cell suspension flows to the sample outlet of the cell capturing and screening chip;
(3) and (4) flowing out the screening cells or cell complexes from the sample outlet.
2. The method for capturing and screening cells according to claim 1, wherein the step (2) comprises reacting at least one pre-immobilized substance in the flow channel wall or/and the cavity during the flowing process until the reacted cell suspension flows to the outlet of the cell capturing and screening chip.
3. The method of claim 1, wherein at least two of said cell capture and selection chips are used, and two adjacent cell capture and selection chips are connected in series.
4. The method for capturing and screening cells according to claim 2, wherein the step (1) is controlled by a matched detection instrument or a pipette is used to add the cell suspension to be screened to the sample inlet.
5. The method of claim 2, wherein the cell capturing and screening chip comprises a sample inlet, a sample outlet, and a flow channel, the sample inlet and the sample outlet are connected through the flow channel, and the flow channel has a plurality of corners from the sample inlet to the sample outlet.
6. The method of claim 5, wherein the flow channel between two adjacent corners is linear.
7. The method of claim 6, wherein the flow channels are alternately connected in series with chambers, and the distance of the flow channels between two non-corner adjacent chambers is the same.
8. The method of capturing and screening cells according to claim 5, wherein the flow channel comprises a first main channel, a second main channel and a surface cavity flow channel, one end of the first main channel is connected to the sample inlet, the other end of the first main channel extends in a direction perpendicular to the first main channel toward the sample outlet and is connected to the surface cavity flow channel, the surface cavity flow channel is further connected to an extending end of the second main channel in a direction perpendicular to the second main channel toward the sample inlet, and the other end of the second main channel is connected to the sample outlet.
9. The method for capturing and screening cells according to claim 6, wherein the width of the flow channel is 20-500 μm, the height of the flow channel is 20-500 μm, and the aspect ratio of the flow channel is 1: 1-2: 1; the cell sap carrying capacity of the flow channel is 1-2500 mu L.
10. The method for capturing and screening cells of claim 7, wherein the diameter of the cavity is 1000-6500 μm, and the ratio of the depth of the cavity to the diameter of the cavity is 1: 1-1: 2.
11. The method for capturing and screening cells according to claim 8, wherein the surface cavity flow channel is a hexagonal flow channel, the hexagonal flow channel has an upward protruding pillar inside, and the hexagonal flow channel has an area of 100-1000 mm2(ii) a The height of the hexagonal flow channel is 0.1-2 mm, and the cell liquid carrying capacity of the hexagonal flow channel is 10-2000 mu L.
CN202010055458.2A 2020-01-17 2020-01-17 Cell capturing and screening method Pending CN111172031A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202010055458.2A CN111172031A (en) 2020-01-17 2020-01-17 Cell capturing and screening method

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202010055458.2A CN111172031A (en) 2020-01-17 2020-01-17 Cell capturing and screening method

Publications (1)

Publication Number Publication Date
CN111172031A true CN111172031A (en) 2020-05-19

Family

ID=70652765

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202010055458.2A Pending CN111172031A (en) 2020-01-17 2020-01-17 Cell capturing and screening method

Country Status (1)

Country Link
CN (1) CN111172031A (en)

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106350439A (en) * 2016-11-10 2017-01-25 上海美吉逾华生物医药科技有限公司 Micro-fluidic chip for cell capture and fluorescent staining
CN207347548U (en) * 2017-10-25 2018-05-11 张沐 A kind of micro-fluidic sugar-modified chip for bacterium identification
CN109852530A (en) * 2019-03-29 2019-06-07 中国科学院上海微系统与信息技术研究所 A kind of micro-fluidic chip and its device and method integrating circulating tumor cell capture, cracking and detection of nucleic acids
CN110093254A (en) * 2019-06-06 2019-08-06 上海海洋大学 It is a kind of for fast Acquisition or detect cell micro-fluidic chip and method
CN110669658A (en) * 2019-11-01 2020-01-10 山东风华生物技术有限公司 Cell capturing and screening device

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106350439A (en) * 2016-11-10 2017-01-25 上海美吉逾华生物医药科技有限公司 Micro-fluidic chip for cell capture and fluorescent staining
CN207347548U (en) * 2017-10-25 2018-05-11 张沐 A kind of micro-fluidic sugar-modified chip for bacterium identification
CN109852530A (en) * 2019-03-29 2019-06-07 中国科学院上海微系统与信息技术研究所 A kind of micro-fluidic chip and its device and method integrating circulating tumor cell capture, cracking and detection of nucleic acids
CN110093254A (en) * 2019-06-06 2019-08-06 上海海洋大学 It is a kind of for fast Acquisition or detect cell micro-fluidic chip and method
CN110669658A (en) * 2019-11-01 2020-01-10 山东风华生物技术有限公司 Cell capturing and screening device

Similar Documents

Publication Publication Date Title
CN111295245B (en) Systems and methods for separating and analyzing cells
US7258837B2 (en) Microfluidic device and surface decoration process for solid phase affinity binding assays
JP5663574B2 (en) Microfluidic analysis platform
CN101419167B (en) High capture rate and high-sensitivity microflow control SPR bio-sensing method and device
CN101661038A (en) Special multi-channel micro-flow controller used in syphilis diagnosis and using low-priced conducting material
CN101587124A (en) Micro-fluidic chip special for diagnosing syphilis by aid of organic conductor material technology
CN103103120A (en) Integrated temperature controlled PCR-CE (polymerase chain reaction-capillary electrophoresis) micro-fluidic chip and preparation method thereof
CN113244970B (en) Microfluidic chip for nucleic acid extraction and PCR detection and application thereof
CN113976195B (en) Microfluidic chip for exosome separation and enrichment and exosome surface protein analysis method
CN105723203A (en) Magnetic separation
JP2010531456A (en) Module for detecting specimen in fluid and chip having the module
CN110468026B (en) Microfluidic chip for optical fiber photodynamic cell manipulation
CN111068801A (en) Self-driven micro-fluidic chip
CN107148468A (en) The microfluidic device with smooth surface for being enriched with rare cell and biomarker from biofluid
Wang et al. A microfluidic robot for rare cell sorting based on machine vision identification and multi-step sorting strategy
CN109097264B (en) Microfluidic cell adsorption chip for cell patch clamp
CN113804658B (en) Microfluidic flow channel structure, detection system and use method thereof
CN111172031A (en) Cell capturing and screening method
CN106841591B (en) A kind of test card of capillary power drive
CN112980677A (en) Micro-fluidic chip for analyzing and sorting tumor cell migration capacity and preparation process
CN211865068U (en) Multi-channel micro-fluidic chip
CN211847950U (en) Cell capturing and screening device
CN108949497B (en) Specific single cell fixed-point capturing chip for trace circulating tumor cells
CN110669658A (en) Cell capturing and screening device
CN109939751B (en) Microfluidic chip, detection device and detection method for whole blood detection

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination