CN103642672B - Biochip for high efficiently separating cells - Google Patents
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- CN103642672B CN103642672B CN201310602809.7A CN201310602809A CN103642672B CN 103642672 B CN103642672 B CN 103642672B CN 201310602809 A CN201310602809 A CN 201310602809A CN 103642672 B CN103642672 B CN 103642672B
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- 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
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Abstract
A biochip for high efficiently separating cells is composed of a cover plate layer and a slide glass arranged below the cover plate layer; a micro-channel which is used for high efficiently separating cells is arranged in the cover plate layer, the micro-channel comprises a fluid inlet, a separating area, and fluid outlets, which are orderly communicated; a first separating area, which communicates with the separating area and the fluid inlet, is composed of a plurality of alternatively arranged expanding sections and a symmetric sharp corner structure; a second separating area is composed of three channels, which are the branches of the first separating area, the fluid outlets comprises a middle fluid outlet and two lateral fluid outlets, and the three fluid outlets respectively communicate with the three channels in the second separating area; the fluid inlet and the fluid outlets are through holes which are arranged in the cover plate layer, the separating areas are typhlosoles which are arranged in the cover plate layer, and the bottom part communicates with the surface of the slide glass. The cells with a large size have a large inertia, so the cells can quickly move to the center of the micro-channel and are discharged from the middle fluid outlet, the cells with a small size are arranged along the two sides of the micro-channel and discharged from the two lateral fluid outlets, so that the high efficient separation of cells with different sizes is achieved. The biochip can be widely applied to the fields of bio-medicine and environment supervision.
Description
Technical field
The present invention relates to a kind of biochip, be specifically related to a kind of biochip for cell high-efficient separation.
Background technology
In medical field, blood testing is an important channel of assessment human health status.Hemocyte in blood is mainly divided into red corpuscle, white corpuscle and thrombocyte.Red corpuscle is generally double intended circle plate-like, in human body, mainly play the effect that material is carried, as all relying on red corpuscle transportation, the nutritive substance of the oxygen sucking from lung and digestive tube absorption could arrive each tissue of whole body, the carbonic acid gas that simultaneously tissue metabolism produces and other refuses also rely on red corpuscle in blood and are transported to lung, kidney etc. and locate excretion, thus the eubolism of maintenance health.White corpuscle is the colourless spherule cell that has core, generally large compared with erythrocyte volume, can do amoeboid movement and enter surrounding tissue through capillary vessel, bring into play its defence and immunologic function, this is because white corpuscle can be engulfed foreign matter generation antibody, treats, resists cause of disease invasion and the immunology of disease is played an important role at body injury.Under light microscopic, according to having or not specific granule can be divided into granulocyte and nongranular leucocyte two classes in white corpuscle matter, granulocyte can, according to the look of having a liking for of particle, be divided into neutrophil leucocyte, eosinophilic granulocyte and basophilic granulocyte again.Nongranular leucocyte has two kinds, monocyte and lymphocyte.Five kinds of cells in white corpuscle are had one's own work to do: neutrophil leucocyte surrounds and kill bacteria and foreign matter in human body wound, and it also can engulf the non-viable non-apoptotic cell in body in body health is safeguarded, participate in inflammatory reaction; Acidophilic granule cell can be eliminated the toxicity of foreign matter; Basophilic granulocyte can discharge anti-coagulant and distends the blood vessels and shrink; Lymphocyte can make tumour dwindle.Wherein T lymphocyte mainly participates in cellular immunization, and bone-marrow-derived lymphocyte mainly participates in humoral immunization.In addition, lymphocyte also synthesizes and secretes multiple lymphokine and participates in hematopoietic regulation; Monocyte can be removed, kill external microorganism, eliminates injured or dead cell and cell debris.Thrombocyte plays an important role in hemostasis, and in normal physiological situation, thrombocyte has metastable quantity.When the platelet content in blood changes, patient's hemostasis and coagulation function will occur extremely.Meeting crisis patient's life security when serious.By detecting hematoblastic content in blood, correct evaluating patient hemostasis and coagulation function are had great significance.Detected result can be used as the reference index whether patient needs to carry out platelet transfusion simultaneously.
Therefore all kinds of hemocytes in blood are had one's own work to do, and HUMAN HEALTH is played a different role, and detect exactly in blood content and the attribute of certain or multiple hemocyte, and assessment human health status is had great significance.In order to detect exactly content and the attribute of certain hemocyte in blood, or certain cell is cultivated and research, target cell and other cellular segregation need to be opened, to get rid of the impact of other cells.In addition, in fields such as medical diagnosis on disease, environmental monitorings, also need cellular segregation.As needed T lymphocyte to separate in other cells from blood in the diagnosis of acquired immune deficiency syndrome (AIDS) and research; The diagnosis of malaria and treatment need to be separated infected red corpuscle from normoerythrocyte; The monitoring of foetus health situation need to be separated the red corpuscle of fetus from parent periphery blood; Alga cells in water body is carried out to separation and detection, and prevention and the improvement of for water, polluting provide direct reference.
Traditional hemocyte separation method mainly comprises centrifugal cell separation technology, fluidic cell sorting technology, electrophoresis-type cell separation technology.Centrifugal cell separation technology is the density variation based on different cells, uses whizzer that target cell is separated.Fluidic cell sorting technology is to make to wrap in the nozzle that the cell in sheath fluid is controlled by HF oscillation, the drop that formation comprises individual cells, then use laser beam to be irradiated to successively on each cell, different cells can send different scattered lights and fluorescence, by detection equipment, detect the optical signalling that each cell sends, then convert electrical signal to and input again computer processing, to judge the kind of cell, and then utilize certain means that target cell is separated from other cells.Electrophoresis-type cell separation technology is the difference of utilizing the surface potential of different cells, and under electric field action, different cells can produce different mobility speeds, according to the difference of cell mobility speed, target cell is separated.Though traditional cell isolation method can be separated target cell from other cells, but there is following shortcoming: 1) expensive, cost is high, is unfavorable for popularization and use in basic medical unit; 2) need additionaling power, consume energy high; 3) bulky, be not easy to carry; 4) complex structure, needs precision equipment auxiliary; 5) use and maintenance expense is high.
Summary of the invention
For solving above-mentioned problems of the prior art, the object of the present invention is to provide a kind of biochip for cell high-efficient separation, have integrated, precision is high, simple in structure, operate simple and easyly, cost is low, the features such as volume is little, will play a significant role in fields such as biomedicine, environmental monitoring and mechanical detection.
In order to achieve the above object, the present invention adopts following technical scheme:
For a biochip for cell high-efficient separation, biochip is comprised of cover plate layer 4 and the slide glass 5 being placed under cover plate layer 4; In described cover plate layer 4, offer the microchannel for cell high-efficient separation, described microchannel comprises liquid adding hole 1, disengaging zone 2 and the leakage fluid dram 3 being communicated with successively, the first disengaging zone 2-1 that described disengaging zone 2 and liquid adding hole 1 are communicated with is comprised of a plurality of expansion segments that distribute alternately and symmetrical horn structure, the the second disengaging zone 2-2 being communicated with leakage fluid dram 3 is three passages by the first disengaging zone 2-1 branch, and described leakage fluid dram 3 comprises middle leakage fluid dram 3-1 and the both sides leakage fluid dram 3-2 that three passages with the second disengaging zone 2-2 are communicated with respectively; Described liquid adding hole 1 and the through hole of leakage fluid dram 3 for offering on cover plate layer 4, the sidewalk for visually impaired people of described disengaging zone 2 for offering on cover plate layer 4, bottom and slide glass 5 surfaces communicate.
Described microchannel is positioned at the combine middle position at place of cover plate layer 4 and slide glass 5.
Described cover plate layer 4 and slide glass 5 combine by the irreversible processing of plasma.
Described the first disengaging zone 2-1 is comprised of 100 groups of symmetrical horn structure and expansion segment.
Described liquid adding hole 1 and leakage fluid dram 3 are cylindrical hole.
The degree of depth of described liquid adding hole 1 and leakage fluid dram 3 is 50 μ m, and diameter is 10mm.
The wedge angle angle of described symmetrical horn structure is 45 degree, the length that the wedge angle of symmetrical horn structure stretches into disengaging zone 2 is 80 μ m, length on wedge angle streamwise is 80 μ m, two relative wedge angles are at a distance of 40 μ m, the distance of adjacent wedge angle on flow direction is 200 μ m, and expansion segment width is 200 μ m.
The material of described cover plate layer 4 is polymetylmethacrylate or polydimethylsiloxane.
The material of described slide glass 5 is glass or silicon.
The present invention is based on different cells and there is different inertia.The cell that size is large has larger inertia, when stream of cells is during through microchannel of the present invention, is subject to inertia lift and centrifugal force simultaneously.Inertia lift is shifted cell onto the both sides of microchannel, and the centrifugal force being caused by horn structure is pushed into cell the center of passage.Maxicell, owing to having larger inertia, under the effect of inertia lift and centrifugal force, will move to the center of microchannel quickly than minicell, last maxicell is got rid of by middle leakage fluid dram, and minicell enters both sides leakage fluid dram, realizes the separation of different cells.
Compared to the prior art, tool has the following advantages in the present invention:
1) cost is low, cheap, compares with other tradition and active microfluid cell separation technologies, and the present invention is without expensive utility appliance, thereby commercial marketing is offered convenience.
2) volume is little, is easy to carry, and can be conveniently used in the aspects such as domestic medicine and individual health care.
3) precision is high, can realize the complete separation of different size cell, compares with the microfluid cell separation technology of other passive types, can under the effect without external force field, realize the complete separation of different size cell, has improved the precision of cellular segregation.
4) physical construction is simple, processing ease, and processing and making process is simple and convenient.
5) integrated, extensibility is high, can combine with the biochip with other functions, promotes the integrated and automatization of analytical system.
Accompanying drawing explanation
Fig. 1 is the vertical view of cell high-efficient separating bio chip of the present invention.
Fig. 2 be Fig. 1 along A-A to sectional view.
Fig. 3 be Fig. 1 along B-B to sectional view.
Fig. 4 is the schematic three dimensional views of cell high-efficient separating bio chip of the present invention.
Fig. 5 is cell high-efficient separating bio chip particle experiment results figure of the present invention.
Embodiment
Below in conjunction with the drawings and specific embodiments, the present invention is further detailed explanation.
As shown in Figures 1 to 4, a kind of biochip for cell high-efficient separation of the present invention, this biochip is processed by standard soft lithography, cover plate layer 4 and the slide glass 5 being placed under cover plate layer 4, consists of; In cover plate layer 4, offer the microchannel for cell high-efficient separation, microchannel comprises liquid adding hole 1, disengaging zone 2 and the leakage fluid dram 3 being communicated with successively, the first disengaging zone 2-1 that disengaging zone 2 and liquid adding hole 1 are communicated with is comprised of a plurality of expansion segments that distribute alternately and symmetrical horn structure, the the second disengaging zone 2-2 being communicated with leakage fluid dram 3 is three passages by the first disengaging zone 2-1 branch, and leakage fluid dram 3 comprises middle leakage fluid dram 3-1 and the both sides leakage fluid dram 3-2 that three passages with the second disengaging zone 2-2 are communicated with respectively; Liquid adding hole 1 and the through hole of leakage fluid dram 3 for offering on cover plate layer 4, the sidewalk for visually impaired people of disengaging zone 2 for offering on cover plate layer 4, bottom and slide glass 5 surfaces communicate.
As the preferred embodiment of the present invention, described microchannel is positioned at the combine middle position at place of cover plate layer 4 and slide glass 5.
As the preferred embodiment of the present invention, described cover plate layer 4 and slide glass 5 combine by the irreversible processing of plasma.
As the preferred embodiment of the present invention, described the first disengaging zone 2-1 is comprised of 100 groups of symmetrical horn structure and expansion segment.
As the preferred embodiment of the present invention, described liquid adding hole 1 and leakage fluid dram 3 are cylindrical hole.
As the preferred embodiment of the present invention, the degree of depth of described liquid adding hole 1 and leakage fluid dram 3 is 50 μ m, and diameter is 10mm.
As the preferred embodiment of the present invention, the wedge angle angle of described symmetrical horn structure is 45 degree, the length that the wedge angle of symmetrical horn structure stretches into disengaging zone 2 is 80 μ m, length on wedge angle streamwise is 80 μ m, two relative wedge angles are at a distance of 40 μ m, the distance of adjacent wedge angle on flow direction is 200 μ m, and expansion segment width is 200 μ m.
As the preferred embodiment of the present invention, the material of described cover plate layer 4 is polymetylmethacrylate or polydimethylsiloxane.
As the preferred embodiment of the present invention, the material of described slide glass 5 is glass or silicon.
Principle of work of the present invention is: cell sample liquid is annotated into microchannel by liquid adding hole 1, then enters the disengaging zone 2 for cellular segregation, and cell is subject to inertia lift and centrifugal force in disengaging zone 2 simultaneously.Larger-size cell has larger inertia, move to quickly the center of microchannel, then by middle leakage fluid dram 3-1, discharged, the cell that size is less is arranged in the both sides of microchannel in disengaging zone 2, finally by both sides leakage fluid dram 3-2, discharged, realize the high efficiency separation of the cell varying in size.
So that being separated into example, the red corpuscle in blood and white corpuscle illustrate that specific embodiments of the invention are as follows below:
1. extract pending blood sample;
2. under the driving of driving arrangement (as syringe pump), blood sample is filled in biochip of the present invention by adding liquid zone 1 with certain flow;
3. the centrifugal force that the red corpuscle in blood and white corpuscle are subject to inertia lift simultaneously and are caused by horn structure in disengaging zone 2.The more erythrocytic volume of white corpuscle is larger, has larger inertia.Under the acting in conjunction of two kinds of power, in disengaging zone 2, white corpuscle will earlier move to the center of microchannel compared with red corpuscle, and red cell distribution is in the both sides of microchannel;
4. behind disengaging zone 2, white corpuscle enters middle leakage fluid dram 3-1; Red corpuscle enters both sides leakage fluid dram 3-2;
5. from middle leakage fluid dram 3-1, extract white corpuscle, from the leakage fluid dram 3-2 of both sides, extract red corpuscle, for next step research.
For the reliability of checking separating effect of the present invention, the fluorescence granules of polystyrene (size corresponds respectively to red corpuscle and white corpuscle) that uses the present invention to be 7.32 microns and 15.5 microns to diameter has carried out separating experiment.Particle is dissolved in deionized water, and adopts the syringe pump that can accurately control flow to inject particle liquid, utilize fluorescent microscope and CCD camera track up particle swarm track.The separating resulting of two kinds of particles as shown in Figure 5, experimental verification the present invention can realize the high efficiency separation of different size particles, thereby in fields such as biomedicine, environmental monitorings, play a significant role.
Claims (8)
1. for a biochip for cell high-efficient separation, it is characterized in that: by cover plate layer (4) and the slide glass (5) that is placed under cover plate layer (4), formed; in described cover plate layer (4), offer the microchannel for cell high-efficient separation, described microchannel comprises the liquid adding hole (1) being communicated with successively, disengaging zone (2) and leakage fluid dram (3), the first disengaging zone (2-1) that described disengaging zone (2) and liquid adding hole (1) are communicated with is comprised of a plurality of expansion segments that distribute alternately and symmetrical horn structure, and the second disengaging zone (2-2) that leakage fluid dram (3) is communicated with is three passages by the first disengaging zone (2-1) branch, described leakage fluid dram (3) comprises middle leakage fluid dram (3-1) and the both sides leakage fluid dram (3-2) that three passages with the second disengaging zone (2-2) are communicated with respectively, described liquid adding hole (1) and leakage fluid dram (3) through hole for offering on cover plate layer (4), described disengaging zone (2) sidewalk for visually impaired people for offering on cover plate layer (4), bottom and slide glass (5) surface communicates, the wedge angle angle of described symmetrical horn structure is 45 degree, the length that the wedge angle of symmetrical horn structure stretches into disengaging zone (2) is 80 μ m, length on wedge angle streamwise is 80 μ m, two relative wedge angles are at a distance of 40 μ m, the distance of adjacent wedge angle on flow direction is 200 μ m, and expansion segment width is 200 μ m.
2. a kind of biochip for cell high-efficient separation according to claim 1, is characterized in that: described microchannel is positioned at the combine middle position at place of cover plate layer (4) and slide glass (5).
3. a kind of biochip for cell high-efficient separation according to claim 1, is characterized in that: described cover plate layer (4) and slide glass (5) combine by the irreversible processing of plasma.
4. a kind of biochip for cell high-efficient separation according to claim 1, is characterized in that: described the first disengaging zone (2-1) is comprised of 100 groups of symmetrical horn structure and expansion segment.
5. a kind of biochip for cell high-efficient separation according to claim 1, is characterized in that: described liquid adding hole (1) and leakage fluid dram (3) are cylindrical hole.
6. a kind of biochip for cell high-efficient separation according to claim 5, is characterized in that: the degree of depth of described liquid adding hole (1) and leakage fluid dram (3) is 50 μ m, and diameter is 10mm.
7. a kind of biochip for cell high-efficient separation according to claim 1, is characterized in that: the material of described cover plate layer (4) is polymetylmethacrylate or polydimethylsiloxane.
8. a kind of biochip for cell high-efficient separation according to claim 1, is characterized in that: the material of described slide glass (5) is glass or silicon.
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| CN201310602809.7A CN103642672B (en) | 2013-11-21 | 2013-11-21 | Biochip for high efficiently separating cells |
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| CN106215985B (en) * | 2016-07-26 | 2018-08-21 | 西安交通大学 | A kind of micro-fluidic chip for quickly mixing and detecting for fluid |
| CN110935238A (en) * | 2019-12-05 | 2020-03-31 | 西安交通大学 | Micro-nano particle enrichment device based on coupling of variable cross-section microchannel and viscoelastic fluid |
| CN113186162B (en) * | 2021-04-19 | 2023-08-15 | 西安交通大学 | Three-dimensional active brain tissue biochip with bionic structure |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1635146A (en) * | 2004-09-23 | 2005-07-06 | 湖南大学 | One-dimensional biological chip and application in gene, protein expression analysis |
| CN101765762A (en) * | 2007-04-16 | 2010-06-30 | 通用医疗公司以马萨诸塞州通用医疗公司名义经营 | Systems and methods for particle focusing in microchannels |
| TW201132757A (en) * | 2010-03-16 | 2011-10-01 | Nat Univ Tsing Hua | Microfluidic chip and method using the same |
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| US20080067068A1 (en) * | 2006-09-19 | 2008-03-20 | Vanderbilt University | DC-dielectrophoresis microfluidic apparatus, and applications of same |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1635146A (en) * | 2004-09-23 | 2005-07-06 | 湖南大学 | One-dimensional biological chip and application in gene, protein expression analysis |
| CN101765762A (en) * | 2007-04-16 | 2010-06-30 | 通用医疗公司以马萨诸塞州通用医疗公司名义经营 | Systems and methods for particle focusing in microchannels |
| TW201132757A (en) * | 2010-03-16 | 2011-10-01 | Nat Univ Tsing Hua | Microfluidic chip and method using the same |
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