CN207749127U - A kind of metabolism detection chip of individual cell level - Google Patents
A kind of metabolism detection chip of individual cell level Download PDFInfo
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- CN207749127U CN207749127U CN201721668616.1U CN201721668616U CN207749127U CN 207749127 U CN207749127 U CN 207749127U CN 201721668616 U CN201721668616 U CN 201721668616U CN 207749127 U CN207749127 U CN 207749127U
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Abstract
The utility model discloses a kind of metabolism detection chips of individual cell level.The metabolism detection chip of the individual cell level includes more than one culture detection unit, each culture detection unit includes liquid stream layer, pneumatic control layer and breathable elastic film, the breathable elastic film is set between the liquid stream layer and pneumatic control layer, the liquid stream layer includes one or more fluid pipe, at least more than one capturing unit is distributed in a fluid pipe bottom, the pneumatic control layer is at least used to that breathable elastic film to be driven to generate the deformation and/or displacement for tending to liquid stream layer, to make microballoon with dissolved oxygen and/or pH detection functions of at least one entrained by the fluid that breathable elastic film and selected capturing unit coordinate capture to be flowed through out of fluid pipe corresponding with selected capturing unit or unicellular.The detection chip of the utility model can probe into unicellular growth conditions and metabolic condition under different dissolved oxygen gradients, realize the real-time detection to dissolved oxygen and acidity situation of change.
Description
Technical field
The utility model is related to the metabolism detection chips more particularly to one kind of a kind of individual cell level can detect in difference
The chip of single cells grown and metabolic condition under dissolved oxygen gradient, belongs to micro-fluidic chip and biotechnology.
Background technology
Cell is organism structure and the base unit of function.Most of research is often using cell colony as research pair
As obtaining the result is that entire cell mass or the macroscopic token of tissue level.But between the Different Individual of allogenic cell there is
Significant microheterogeneity (heterogeneity), the experimental result based on a large amount of cells are difficult to reflect that the life on individual cell level is lived
Dynamic rule.Therefore there is an urgent need to research and develop to be based on single celled culture and analysis method, for studying the difference between different cell individuals
The opposite sex understands heredity and the metabolic mechanism of cell.It is to carry out unicellular culture and analysis that individual cells are obtained from a large amount of cells
The first step.Traditional unicellular acquisition methods frequently with the method that Macrodilution or micromanipulation are carried out to cell mass come into
Row.Whole operation step complexity is cumbersome, and unicellular acquisition efficiency is low.
Oxygen plays consequence in cell function and behavior.The growth rate of cell and metabolism and albumen
Synthesis greatly depends on the level of oxygen in culture medium.In human body, the level of oxidation is very important.When change oxygen
Content, from it is oxygen-enriched to hypoxemia again to anaerobic, it may occur that strong biological respinse.Some researches show that hypoxemia can promote to regenerate
The proliferation of stem cell.However, hypoxemia can also lead to pathological generation, include generation and the cardiovascular disease of inflammation, tumour
Disease etc..Therefore research single cell growth state and metabolic condition under different dissolved oxygen conditions is extremely important.Currently, in hypoxemia
The method for studying cell behavior in environment has anoxic work station and anoxic cell, but these cannot form oxygen concentration gradients.
In order to meet the above needs, various microfluidic devices have been exploited for single celled capture and stringent control
Amount of oxygen.
Microflow control technique provides outstanding platform to study the microenvironment of dissolved oxygen gradient.There are many researchs thin at present
Behavioral aspect of the born of the same parents under different oxygen concentrations.Such as 2011, it is dense that Chen, etal. report a kind of oxygen for cell culture
Gradient chip is spent, antitumor drug Tirapazamine under anaerobic condition is had detected in the case where cell is in oxygen concentration gradient
(TPZ) to the influence of tumour cell.However, what present technology often studied is the entire group of cell under different oxygen concentrations gradient
Behavior.Therefore, they do not study the heterogeneity of the metabolic characteristics between Different Individual cell in same culture, do not have
Probe into behavior of the individual cells under different oxygen concentrations.It is evident that the experimental result based on a large amount of cells is difficult to reflect list
Vital movement rule on cellular level.
2012, LaimonasKelbauskas, etal. reported a kind of wear rate for studying oxygen under individual cell level
Chip.The chip can detach cell by using microchamber, then utilize non-contacting piezoelectricity liquid make-up machine device people will
Oxygen sensitive dye instills in microchamber, then seals cell, can thus measure the consumption of oxygen under individual cell level.The chip
Although flux is high, oxygen sensitivity detection means is complicated.Further, which reported in 2017 under individual cell level
The situation of change of oxygen and pH, this method discloses the cell subsets that overview is generated with incorrect energy, and can determine to electricity
The cell response changeability of son transfer chain inhibitor and ion uncoupler, the instrument used in same this method costly,
Detection means complex manufacturing process.
In view of this, it is necessary to provide a kind of vital movement rule that can study individual cell level under different dissolved oxygen gradients
The chip of rule, while dissolved oxygen condition is measured in real time.
Utility model content
The main purpose of the utility model is to provide a kind of metabolism detection chips of individual cell level, to overcome existing skill
The deficiency of art.
For realization foregoing purpose, the technical solution adopted in the utility model includes：
The utility model embodiment provides a kind of metabolism detection chip of individual cell level comprising more than one culture
Detection unit, each culture detection unit includes liquid stream layer, pneumatic control layer and breathable elastic film, and the breathable elastic film is set
It is placed between the liquid stream layer and pneumatic control layer, the liquid stream layer includes one or more fluid pipe, wherein at least a liquid stream
More than one capturing unit is distributed in duct bottom, and the pneumatic control layer, which is at least used to that breathable elastic film to be driven to generate, tends to liquid
The deformation and/or displacement of fluid layer, to make breathable elastic film and selected capturing unit coordinate capture from selected capturing unit phase
Answer at least one microballoon or unicellular with dissolved oxygen and/or pH detection functions entrained by the fluid flowed through in fluid pipe.
Compared with prior art, the beneficial effects of the utility model include：
1) the metabolism detection chip reasonable design simple in structure of individual cell level provided by the utility model, first with micro-
Column RAID card residence need study it is unicellular, higher can capture it is unicellular, and capture cell it is not easily to fall off；Together
When, using design different oxygen concentrations gradient generate structure, come probe into the unicellular growth conditions under different dissolved oxygen gradients with
And metabolic condition, while realizing the real-time detection in unicellular incubation to dissolved oxygen and acidity situation of change, it is single for research
Vital movement rule on cellular level provides a kind of scheme；It further can also be used to study unicellular in different dissolved oxygens
Under the conditions of interaction with drug；
2) the metabolism detection chip of individual cell level provided by the utility model be integrated with it is unicellular capture, Dissolved oxygen regulation with
Dissolved oxygen and acidity are detected on one, and manufacture craft is simple, and the element number integrated on unit area can also greatly improve, and have more
High culture assays efficiency.
Description of the drawings
It, below will be to institute in embodiment or description of the prior art in order to illustrate more clearly of the technical solution of the utility model
Need attached drawing to be used simply to be introduced, it should be apparent that, drawings discussed below as just the utility model one
A little embodiments for those of ordinary skill in the art without creative efforts, can also be according to these
Attached drawing obtains other accompanying drawings.
Fig. 1 is the vertical view of the metabolism detection chip of individual cell level in one exemplary embodiments of the utility model.
Fig. 2 be individual cell level in Fig. 1 metabolism detection chip in part A close-up schematic view.
Fig. 3 is the cross-sectional view of B-B along the metabolism detection chip of individual cell level in Fig. 1.
Fig. 4 is the dissolved oxygen gradient formed in the metabolism detection chip of individual cell level in one exemplary embodiments of the utility model
Schematic diagram.
Reference sign：110- liquid stream layers, 1101- fluid pipes, 120- pneumatic control layers, 130- breathable elastic films,
1201,1202,1203,1204- pneumatic control pipelines, 1301- microtrabeculaes, 1001- dissolved oxygens detect microballoon, and 1002-pH detections are micro-
Ball, 1000- are unicellular.
Specific implementation mode
As previously mentioned, in view of the defect of the prior art, inventor is able to propose this through studying for a long period of time and largely putting into practice
The technical solution of utility model, studied required for mainly being blocked first with micro-column structure it is unicellular, then using setting
The different oxygen concentrations gradient of meter generates structure, the unicellular growth under dissolved oxygen gradient and metabolic condition can be successfully probed into, to grind
The vital movement rule studied carefully on individual cell level provides a kind of scheme.It as follows will be to the technical solution, its implementation process and original
Reason etc. is further explained.
As the one side of technical solutions of the utility model, involved by be a kind of metabolism detection of individual cell level
Chip comprising more than one culture detection unit, each culture detection unit include liquid stream layer, pneumatic control layer and breathe freely
Elastic membrane, the breathable elastic film are set between the liquid stream layer and pneumatic control layer, and the liquid stream layer includes one or more
More than one capturing unit is distributed in fluid pipe, wherein at least a fluid pipe bottom, and the pneumatic control layer is at least used for
Breathable elastic film is driven to generate the deformation and/or displacement for tending to liquid stream layer, to make breathable elastic film match with selected capturing unit
It closes at least one with dissolved oxygen and/or pH entrained by the fluid that capture is flowed through out of fluid pipe corresponding to selected capturing unit
The microballoon of detection function is unicellular.
Among an embodiment, the pneumatic control layer includes one or more pneumatic control pipeline, wherein at least a gas
Dynamic control pipeline is intersected with an at least fluid pipe at more than one capture region, when pneumatically being controlled to described at least one
When tubing giving sufficient strength is passed through the gas with setting pressure, at least one pneumatic control pipeline can drive the poromeric elastic
Film generates the deformation and/or displacement of the trend liquid stream layer, the selected capturing unit point at selected capture region
It is distributed in the selected capture region.
Among an embodiment, when being passed through the fluid containing oxygen into an at least pneumatic control pipeline, until
Less at the capture region, the oxygen in an at least pneumatic control pipeline can enter fluid flow tube through breathable elastic film
Road.
Among an embodiment, the capturing unit includes the micro-pillar array being distributed in fluid pipe, the microtrabeculae
Array includes a plurality of microtrabeculaes, wherein three or more adjacent microtrabeculaes are enclosed a capture hole, the capture hole is used for
Capture the microballoon or unicellular.
Among an embodiment, the culture detection unit further includes an at least micro-valve, and each micro-valve includes being distributed in
A pneumatic control pipeline in the pneumatic control layer, the pneumatic control pipeline and a fluid pipe are handed at selected location
Fork, when being passed through gas of the pressure not less than a threshold value to the pneumatic control pipeline, the pneumatic control pipeline can drive
The breathable elastic film generates the deformation and/or displacement of the trend liquid stream layer, to select an at least fluid pipe
It is closed at position, as oxygen concentration gradient generates structure.
Gas in pneumatic control pipeline can pass through bullet of breathing freely in the infall with fluid pipe (for annular closed pipeline)
In property membrane diffusion to the culture solution of annular closed pipeline.By changing the area and friendship that pneumatic control pipeline intersects with fluid pipe
The quantity of crunode can change the amount that gas diffusion enters culture solution, dense to change the dissolved oxygen in culture detection unit
Degree；In addition it can also change pneumatic control pipeline and be passed through the concentration of gas to change the dense of the dissolved oxygen in culture detection unit
Degree.If duct size, shape, gas injection ingredient are completely the same, then identical solubility concentration can be achieved.Pneumatic control pipeline
In gas can be oxygen or oxygen containing gas.It may be implemented by the design of pneumatic control pipeline configuration single in culture detection
Oxygen concentration gradient in member, and then the unicellular metabolism behavior under oxygen concentration gradient can be studied.
Among an embodiment, the culture detection unit further includes an at least Micropump, and each Micropump includes being distributed in
Two or more pneumatic control pipelines in the pneumatic control layer, described two or more pneumatic control pipelines and an at least fluid flow tube
Road intersects at different locations, when in the setting period, into described two or more pneumatic control pipelines, input has different pressures respectively
When the gas of power, described two or more pneumatic control pipelines can drive the different regional areas of the breathable elastic film to produce respectively
Raw different degrees of deformation and/or displacement can drive liquid stream one-way flow to be generated in an at least fluid pipe
Power.
Among an embodiment, capturing unit is uniformly distributed in below corresponding pneumatic control pipeline in the liquid stream layer
And it is connected to the pneumatic control pipeline.
Further, the capturing unit each in culture detection unit is spaced apart in corresponding fluid pipe bottom, for example,
It is intervally arranged with 500 μm, 1500 μm, 2500 μm, 3500 μm.
Among an embodiment, the capture hole includes the hole, molten to capture for capturing pH detection microballoons
Oxygen detects the hole of microballoon and to capture the hole of cell.
Pneumatic control pipeline can be used for dissolved oxygen detection microballoon or pH detection microballoons or single celled when wherein, at the beginning
Capture, pneumatic control pipeline generates under the conditions of pouring certain pressure drives the breathable elastic film to generate different degrees of deformation
And/or displacement, to be formed between the breathable elastic film and liquid stream layer list in the liquid of fluid pipe is flowed through for retaining
A dissolved oxygen detection microballoon or pH detection microballoons or single celled capture region.
Further, to capture cell hole diameter be slightly smaller than or equal to individual cells diameter.
Likewise, slightly smaller than or straight equal to single dissolved oxygen detection microballoon to the diameter for capturing the hole that pH detects microballoon
Diameter.
Likewise, slightly smaller than or straight equal to single pH detection microballoons to the diameter for capturing the hole that dissolved oxygen detects microballoon
Diameter.
Preferably, the microballoon includes pH detection microballoons and dissolved oxygen detection microballoon, and the pH detections microballoon, dissolved oxygen detection are micro-
The diameter of ball and cell is different.
Further, the diameter of the dissolved oxygen detection microballoon is less than diameter of the diameter less than cell of pH detection microballoons.
Further, the material of the dissolved oxygen detection microballoon not only can only comprising oxygen sensitive materials but also can include that oxygen is quick
Ability and the insensitive material of oxygen (doing reference).
Among an embodiment, the dissolved oxygen detection microballoon is synthesized using emulsion polymerization.The utility model is using breast
Liquid polymerization detects microballoon to synthesize dissolved oxygen, and this method synthesizes the efficient of microballoon, implements simple possible.
Among an embodiment, the preparation method of the dissolved oxygen detection microballoon includes：Make comprising oxygen sensitive materials, packet
The hybrid reaction system for wrapping up in material performed polymer, crosslinking agent, surfactant, organic solvent and water carries out polymerizeing instead in 40~80 DEG C
5~10h is answered, the dissolved oxygen detection microballoon is obtained.
Among an embodiment, the preparation method of the dissolved oxygen detection microballoon includes：
There is provided comprising oxygen sensitive materials (being referred to as oxygen sensitivity dyestuff), lapping performed polymer, crosslinking agent and
The homogeneous mixture system of organic solvent uniformly mixes the homogeneous mixture system with the aqueous solution of surfactant, and in 40
5~10h of polymerisation is carried out at~80 DEG C, obtains the dissolved oxygen detection microspheres solution.
Preferably, the mass ratio of the lapping performed polymer and crosslinking agent is 5：1~10：1.
Further, when the synthesis of dissolved oxygen detection microballoon, in addition to that can select that PDMS is used to contain oxygen sensitive materials as package
Material, it is also an option that other replace such as PS, or PDMS is mixed with PS for wrapping up containing oxygen sensitive materials.
Preferably, the material of the dissolved oxygen detection microballoon not only can only comprising oxygen sensitive materials but also can include that oxygen is sensitive
Property material and the insensitive material of oxygen (doing reference).
Further, the oxygen sensitive materials include PtTFPP, PtOEP, PtOEPK etc., but not limited to this.
Further, a concentration of 1~4mmol/L of the oxygen sensitive materials.
Preferably, the insensitive material of the oxygen includes PFO (polyfluorene), MFY (fluorescein) etc., but not limited to this.
Further, surfactant can be selected as SDS (lauryl sodium sulfate), PVA (polyvinyl alcohol) etc., but not
It is limited to this.
It is further preferred that when the surfactant selects SDS, the concentration range of SDS is 0.05~0.2mol/L；
When the surfactant selects PVA, the content of PVA is 1~15wt% in the hybrid reaction system.
Preferably, the crosslinking agent includes silane coupling agent, and further, the crosslinking agent has alkenyl (predominantly
Vinyl) and end have Cl-, NH2, SH-, epoxy group, N3, the functions such as (methyl) acryloxy and isocyanate group
The alkyl of at least one of group, the especially preferably crosslinking agent containing vinyl.
Preferably, the organic solvent includes that dichloromethane, chloroform, tetrahydrofuran, styrene and toluene etc. both may be used
The solvent of the lappings performed polymer such as PDMS can be dissolved to dissolve oxygen sensitive materials again, but not limited to this.
Further, when the synthesis of dissolved oxygen detection microballoon, the size of microballoon can be lived by the amount of adjusting solvent and/or surface
The concentration of property agent controls.Such as when the use scope of solvent be 3~5 milliliters when, synthesis dissolved oxygen detection microspherulite diameter be 8~
15 microns.
Among an embodiment, the preparation method of the dissolved oxygen detection microballoon further includes：With air-distillation, vacuum distillation
Etc. any one of modes the dissolved oxygen detected into the organic solvent in microspheres solution remove.Further, it is preferred to use decompression
The mode of distillation, because the dissolved oxygen detection microballoon dispersibility synthesized in a manner of being evaporated under reduced pressure is preferably, grain size is more uniform.
Wherein, the preparation method of the pH detections microballoon and the preparation method of dissolved oxygen detection microballoon are more similar, diameter
Size can also be adjusted according to the concentration of surfactant and the amount of organic solvent.
Among an embodiment, the shape of the microtrabeculae includes cylindrical type, cuboid, halfpace, cone, open trough
Any one in type structure (meniscus) or two or more combinations, but not limited to this.
Further, a diameter of 10 microns of the microtrabeculae are highly 10 microns.
Among an embodiment, the fluid pipe is set interface on the chip and is in communication with the outside.
Preferably, the fluid pipe includes at least one annular fluid pipe.
Preferably, the material of the breathable elastic film includes dimethyl silicone polymer, but not limited to this.
The metabolism detection chip reasonable design simple in structure of individual cell level provided by the utility model, first with microtrabeculae
RAID card residence need study it is unicellular, higher can capture it is unicellular, and capture cell it is not easily to fall off；Meanwhile
Structure is generated using the different oxygen concentrations gradient of design, to probe into the unicellular growth conditions under different dissolved oxygen gradients and generation
It thanks to situation, while realizing and dissolved oxygen and acidity situation of change are measured in real time in unicellular incubation, it is slender to study
Vital movement rule in born of the same parents' level provides a kind of scheme；It further can also be used to study unicellular in different dissolved oxygen items
Under part with the interaction of drug.
As the other side of technical solutions of the utility model, involved by be aforementioned individual cell level metabolism inspection
Survey the application of chip.
The metabolism detection method of the individual cell level of the utility model includes：
The metabolism detection chip of individual cell level above-mentioned is provided；
Carrier fluid of the injection comprising dissolved oxygen detection microballoon and pH detection microballoons into the liquid stream layer of the chip, and make the load
Liquid is flowed according to predetermined flow rate in selected fluid pipe；
Drive breathable elastic film by the part dissolved oxygen detection microballoon and pH detection microballoons in the carrier fluid with pneumatic control layer
It is squeezed into selected capturing unit, realizes the capture of dissolved oxygen detection microballoon and pH detection microballoons, then removes the selected fluid pipe
The interior dissolved oxygen detection microballoon at large for obtaining unit capture and pH detection microballoons；
Celliferous cell culture fluid is wrapped in injection at least into the selected fluid pipe, then is driven with pneumatic control layer
Unicellular in the cell culture fluid is squeezed into the selected capturing unit by gas elastic membrane, realizes single celled capture；
And
To it is described it is unicellular cultivate, and in incubation and/or after culture, detected with captured dissolved oxygen
Microballoon and pH detection microballoons detect dissolved oxygen content and acidity situation in the single celled cultivating system.
Preferably, the detection method further includes：To it is described it is unicellular proceed by culture when and/or culture process
In, into an at least pneumatic control pipeline for the pneumatic control layer, input includes the fluid of oxygen, and keeps oxygen therein saturating
It crosses breathable elastic film and enters the selected fluid pipe, and then change the oxygen dissolved in the selected fluid pipe.
Further, the oxygen-bearing fluid includes oxygen-containing gas, oxygen-bearing liquid or the chemical reagent for producing oxygen/deoxidation.
Preferably, the oxygen-containing gas can be in oxygen or oxygen containing gas, such as nitrogen and carbon dioxide at least
A kind of combination with oxygen, but not limited to this.
Preferably, the dissolved oxygen detection method further includes：Dissolved oxygen detection microballoon is detected by optical imagery and pH detections are micro-
The light intensity signal that ball generates, to measure the dissolved oxygen content and acidity situation in the single celled cultivating system.
The metabolism detection chip of individual cell level provided by the utility model be integrated with it is unicellular capture, Dissolved oxygen regulation with it is molten
Oxygen and acidity are detected on one, and manufacture craft is simple, and the element number integrated on unit area can also greatly improve, and have higher
Culture assays efficiency.
The following will be combined with the drawings in the embodiments of the present invention, carries out the technical scheme in the embodiment of the utility model
Detailed description, it is clear that the described embodiments are only a part of the embodiments of the utility model, rather than whole implementation
Example.Based on the embodiments of the present invention, those of ordinary skill in the art institute without making creative work
The every other embodiment obtained, shall fall within the protection scope of the present invention.
Please refer to Fig.1 shown in-Fig. 3, the metabolism detection chip of the individual cell level of one exemplary embodiments of the utility model by
Three structure sheaf compositions stacked on top of one another.Bottom is liquid stream layer 110, and upper layer is pneumatic control layer 120, there is ventilative bullet between two layers
Property film 130 is separated by.There is fluid pipe 1101 (that is, annular closed pipeline) in liquid stream layer 110, width is 100 microns, a height of 20
Microns, both ends are connected to outer, have 6 raised microtrabeculaes 1301, diameter in 1101 regional area bottom of fluid pipe
It is 10 microns, a height of 10 microns, wherein the region surrounded between intermediate four microtrabeculaes is to capture the hole of cell, on one side one
A microtrabeculae and surround to capture cell hole microtrabeculae between gap be to capture dissolved oxygen detect microballoon hole, separately
On one side a microtrabeculae and surround to capture cell hole microtrabeculae between gap be to capture pH detect microballoon sky
Cave.There is pneumatic control pipeline 1201,1202,1203,1204 in pneumatic control layer 120.Wherein, it is distributed in the pneumatic control layer
Pneumatic control pipeline 1204 be also referred to as steam line, pneumatic control pipeline 1201 be cycle driving pipeline, to be formed
Micropump, pneumatic control pipeline 1202 and 1203 is forming micro-valve.Wherein, steam line is one or several with extraneous air source phase
Pipeline even, all pipeline depths are 10 microns.
The testing principle of the metabolism detection chip of the individual cell level of the utility model is as follows：
Three microtrabeculaes surround a region equal or smaller with dissolved oxygen detection microballoon, i.e., detect microballoon to capture dissolved oxygen
Hole.When dissolved oxygen detects microballoon when this region vertical direction flows through, periodically pressurization makes pneumatic control pipeline 1204 repeatedly
Breathable elastic film generates deformation, and fluid pipe 1101 is periodically squeezed to offside, so has dissolved oxygen detection microballoon to flow through positioned at molten
When oxygen detects the microballoon capture corresponding region in hole, dissolved oxygen detection microballoon is the capture hole being extruded between being stuck in microtrabeculae, complete
The capture of microballoon is detected at single dissolved oxygen.The acquisition mode that pH detects microballoon is similar to above-mentioned dissolved oxygen detection microballoon, and pH is detected
Microballoon is more than dissolved oxygen and detects microballoon.The optical signalling that the single dissolved oxygen detection microballoon and pH detection microballoons of capture generate is by ambient light
Learn detector collect, so as to in fluid pipeline 1101 dissolved oxygen and acidity level be detected.Four microtrabeculaes surround one
A region equal or smaller with cell size, i.e., to capture the hole of cell, when cell is flowed through in this region vertical direction
When, periodically pressurization makes breathable elastic film generate deformation to pneumatic control pipeline 1204 repeatedly, and fluid flow tube is periodically squeezed to offside
Road 1101, when so having cell to flow through region corresponding positioned at cell capture hole, cell is extruded and is stuck between microtrabeculae
Hole is captured, single celled capture is completed.After unwanted cell flushes out pipeline in fluid pipe, this section of fluid pipe contains
Individual cells.When changing liquid, it is only necessary to there is single celled fluid pipe to inject new liquid to capture.In pneumatic control pipeline 1204
Gas can be diffused into fluid pipe in the infall with fluid pipe 1101 (annular closed pipeline) by breathable elastic film 130
In the culture solution of 1101 (annular closed pipelines).The area intersected with fluid pipe 1101 by changing pneumatic control pipeline 1204
The amount that gas diffusion enters culture solution can be changed with the quantity in crosspoint, to change the dissolved oxygen in culture detection unit
Concentration；In addition it can also change pneumatic control pipeline 1204 and be passed through the concentration of gas to change the dissolved oxygen in culture detection unit
Concentration.If duct size, shape, gas injection ingredient are completely the same, then identical solubility concentration can be achieved.Pneumatic control
Gas in pipeline 1204 can be oxygen or oxygen containing gas.It may be implemented by the design of 1204 structure of pneumatic control pipeline
Oxygen concentration gradient in cultivating detection unit, and then the unicellular metabolism behavior under oxygen concentration gradient can be studied.
Wherein, as one, more specifically one of case study on implementation, the metabolism of the individual cell level of the utility model detect core
The application detection process of piece is as described below：
In conjunction with Fig. 1-Fig. 3, first into fluid pipe 1101, injection detects microballoon comprising dissolved oxygen detection microballoon 1001 and pH
1002 mixed solution, be then periodically filled with high-pressure fluid into pneumatic control pipeline 1204, hole capture to be captured list
After a dissolved oxygen detection microballoon 1001 and pH detection microballoons 1002, the air pressure in pneumatic control pipeline 1204 is removed, to fluid pipe
In inject cleaning solution in 1101, extra the dissolved oxygen detection microballoon not captured or pH detection microballoons are gone out, then captured
Single dissolved oxygen detection microballoon and pH detect microballoon.Then cell suspension is injected into fluid pipe 1101, then to pneumatic control
It is periodically filled with high-pressure fluid in pipeline 1204, after hole capture to be captured unicellular 1000, removes pneumatic control pipeline
Air pressure in 1204 injects cleaning solution into fluid pipe 1101, and the extra cell not captured is gone out, then is captured
Unicellular 1000.Then it is filled with oxygenous gas into pneumatic control pipeline 1204, pneumatic control pipeline can be closed after inflation
The interface at 1204 both ends keeps normal pressure, can also continue to rush to pneumatic control pipeline 1204 with lower pressure in incubation
Enter the oxygen-containing gas of moistening, realizes that cell is cultivated in the fluid pipe of different oxygen concentrations gradient, and in its incubation
Dissolved oxygen and acidity situation are measured in real time.
The more typical embodiment of the one of the utility model may include that process is implemented as follows：
One, the synthesis of dissolved oxygen detection microballoon：Above-mentioned microballoon is synthesized using the method for emulsion polymerization.
Experimentation is as follows：
A) surfactant solution for preparing 0.2MolSDS and 14wt%PVA, respectively takes 15ml to be added in the flask of 100ml,
It adjusts temperature adjustment knob and is warming up to 40 degree, rotating speed is adjusted to 300rpm or so.
B) centrifuge tube is taken, the PDMS performed polymers mixed and crosslinking agent 0.6g (curing ratios 10 is added：1).
C) centrifuge tube is taken again, by oxygen sensitive dye PtTFPP and the insensitive dyestuff PFO of oxygen with mass ratio 4：2 ratio
It is dissolved into the chloroform of 4ml, the ultimate density of PtTFPP is 1mmol/l.
D) it b) and c) is mixed above-mentioned, shakes 5min, its substantially uniformity is made to mix.
E) rate of 10 μ l/min of above-mentioned solution d) prepared is instilled in flask with syringe pump.
F) then heat preservation rotation dispersion 2h opens vacuum pump switch and is decompressed to 0.08Mpa or so, after decompression process,
Keep the temperature 1h, you can obtain the suspension of dissolved oxygen detection microballoon.
Two, pH detects the synthesis of microballoon：Above-mentioned microballoon is synthesized using the method for emulsion polymerization.
Experimentation is as follows：
A) surfactant solution for preparing 0.2MolSDS and 8wt%PVA, respectively takes 15ml to be added in the flask of 100ml,
It adjusts temperature adjustment knob and is warming up to 40 degree, rotating speed is adjusted to 300rpm or so.
B) centrifuge tube is taken, the PDMS performed polymers mixed and crosslinking agent 0.6g (curing ratios 10 is added：1).
C) centrifuge tube is taken again, with mHPTS：MPFO=1：1 ratio is dissolved into the chloroform of 4ml.
D) it b) and c) is mixed above-mentioned, shakes 5min, its substantially uniformity is made to mix.
E) rate of 10 μ l/min of above-mentioned solution d) prepared is instilled in flask with syringe pump.
F) then heat preservation rotation dispersion 2h opens vacuum pump switch and is decompressed to 0.08Mpa or so, after decompression process,
Keep the temperature 1h, you can obtain the suspension of pH detection microballoons.
Inventor is carried out referring also to aforementioned preparation process with the other raw materials and process conditions listed in this specification
The preparation of dissolved oxygen detection microballoon, pH detection microballoons.
Three, dissolved oxygen detection microballoon and pH detection microballoons are passed through chip.
The dissolved oxygen of synthesis is detected into microballoon and pH detects microballoon according to 1：It is diluted to proper level after 1 ratio mixing, to
It is passed through above-mentioned detection microballoon mixed solution in fluid pipe 1101 in each culture detection unit, waits for that dissolved oxygen detects 1001 He of microballoon
PH detection microballoons 1002 be trapped in the capture array of lower layer after as shown in figure 3, with water by dissolved oxygen extra in fluid pipe
Detection microballoon or pH detection microballoons wash away.It is finally continuously passed through the air of moistening into pneumatic control pipeline 1204, is examined in culture
The dissolved oxygen gradient formed such as Fig. 4 is surveyed in the fluid pipe of unit.Finally in the fluid pipe 1101 into each culture detection unit
It is passed through celliferous culture solution, carries out the cell culture under dissolved oxygen gradient.Detection and the acid of dissolved oxygen are carried out after culture a period of time
The detection of degree, is detected by optical imagery, collects light intensity signal, carries out the metabolism in incubation under dissolved oxygen gradient accordingly
It measures.Measurement result is shown, by the aforementioned metabolic detection chip of the utility model, can accurately, efficiently be realized unicellular
The measurement of metabolism under horizontal dissolved oxygen gradient.
In conclusion by the above-mentioned technical proposal of the utility model, the metabolism of the individual cell level of the utility model is examined
Unicellular growth conditions and metabolic condition under different dissolved oxygen gradients can be probed by surveying chip, realized and become to dissolved oxygen and acidity
The real-time detection for changing situation, can also be used to study it is unicellular under different dissolved oxygen conditions with the interaction of drug.
It should be noted that herein, the terms "include", "comprise" or its any other variant are intended to non-row
His property includes, so that the process, method, article or equipment including a series of elements includes not only those elements, and
And further include other elements that are not explicitly listed, or further include for this process, method, article or equipment institute it is intrinsic
Element.In the absence of more restrictions, the element limited by sentence "including a ...", it is not excluded that including institute
State in the process, method, article or equipment of element that there is also other identical elements.
It should be appreciated that above-described embodiment is only the technical concepts and features for illustrating the utility model, it is ripe its object is to allow
The personage for knowing technique can understand the content of the utility model and implement according to this, can not limit the utility model with this
Protection domain.It is all according to equivalent change or modification made by the spirit of the present invention essence, should all cover in the utility model
Within protection domain.
Claims (10)
1. a kind of metabolism detection chip of individual cell level, it is characterised in that including more than one culture detection unit, Mei Yipei
Foster detection unit includes liquid stream layer, pneumatic control layer and breathable elastic film, and the breathable elastic film is set to the liquid stream layer
Between pneumatic control layer, the liquid stream layer includes one or more fluid pipe, and wherein at least a fluid pipe bottom is distributed with
More than one capturing unit, the pneumatic control layer at least be used for drive breathable elastic film generate tend to liquid stream layer deformation and/
Or displacement, to make breathable elastic film coordinate capture to be flowed out of fluid pipe corresponding with selected capturing unit to selected capturing unit
At least one microballoon or unicellular with dissolved oxygen and/or pH detection functions entrained by the fluid of warp.
2. the metabolism detection chip of individual cell level according to claim 1, it is characterised in that：The pneumatic control layer packet
One or more pneumatic control pipeline is included, a wherein at least pneumatic control pipeline is with an at least fluid pipe in more than one capture
At region intersect, when at least one pneumatic control pipeline be passed through with setting pressure gas when, it is described at least
One pneumatic control pipeline can drive the breathable elastic film to generate the trend liquid stream layer at selected capture region
Deformation and/or displacement, the selected capturing unit is distributed in the selected capture region.
3. the metabolism detection chip of individual cell level according to claim 2, it is characterised in that：To an at least gas
When being passed through the fluid containing oxygen in dynamic control pipeline, at least at the capture region, an at least pneumatic control pipeline
Interior oxygen can enter fluid pipe through breathable elastic film.
4. the metabolism detection chip of individual cell level according to any one of claim 1-3, it is characterised in that：It is described to catch
It includes the micro-pillar array being distributed in fluid pipe to obtain unit, and the micro-pillar array includes a plurality of microtrabeculaes, wherein adjacent three
A above microtrabeculae is enclosed a capture hole, and the capture hole is used to capture the microballoon or unicellular.
5. the metabolism detection chip of individual cell level according to any one of claim 1-3, it is characterised in that：The training
Foster detection unit further includes an at least micro-valve, and each micro-valve includes the pneumatic control pipe being distributed in the pneumatic control layer
Road, the pneumatic control pipeline intersect with a fluid pipe at selected location, when being passed through pressure to the pneumatic control pipeline
Not less than a threshold value gas when, the pneumatic control pipeline can drive the breathable elastic film to generate the trend liquid stream
The deformation and/or displacement of layer, to will at least a fluid pipe be closed at selected location.
6. the metabolism detection chip of individual cell level according to any one of claim 1-3, it is characterised in that：The training
Foster detection unit further includes an at least Micropump, and each Micropump includes two or more pneumatic controls being distributed in the pneumatic control layer
Tubing giving sufficient strength, described two or more pneumatic control pipelines intersect at different locations with an at least fluid pipe, when in the setting period
When into described two or more pneumatic control pipelines, input has the gas of different pressures respectively, described two or more pneumatic controls
Pipeline can drive the different regional areas of the breathable elastic film to generate different degrees of deformation and/or displacement respectively, to
The power of liquid stream one-way flow can be driven by being generated in an at least fluid pipe.
7. the metabolism detection chip of individual cell level according to claim 4, it is characterised in that：The capture hole includes
The hole of microballoon is detected to capture pH, detects the hole of microballoon and to capture dissolved oxygen to capture the hole of cell.
8. the metabolism detection chip of individual cell level according to claim 4, it is characterised in that：The microballoon includes pH inspections
Micrometer ball and dissolved oxygen detect microballoon, and the diameter of the pH detections microballoon, dissolved oxygen detection microballoon and cell is different.
9. the metabolism detection chip of individual cell level according to claim 4, it is characterised in that：The shape packet of the microtrabeculae
Include any one in cylindrical type, cuboid, halfpace, cone, open trench structure or two or more combinations.
10. the metabolism detection chip of individual cell level according to claim 1, it is characterised in that：The fluid pipe
The interface being set on the chip is in communication with the outside, and the fluid pipe includes at least one annular fluid pipe.
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| CN201721668616.1U CN207749127U (en) | 2017-12-05 | 2017-12-05 | A kind of metabolism detection chip of individual cell level |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109868219A (en) * | 2017-12-05 | 2019-06-11 | 中国科学院苏州纳米技术与纳米仿生研究所 | A kind of the metabolism detection chip and its application of individual cell level |
| CN111378576A (en) * | 2018-12-28 | 2020-07-07 | 上海大学 | Intestinal tract simulation chip and application thereof |
| CN113101992A (en) * | 2021-05-08 | 2021-07-13 | 苏州珀罗汀生物技术有限公司 | Protein synthesis screening method |
-
2017
- 2017-12-05 CN CN201721668616.1U patent/CN207749127U/en active Active
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109868219A (en) * | 2017-12-05 | 2019-06-11 | 中国科学院苏州纳米技术与纳米仿生研究所 | A kind of the metabolism detection chip and its application of individual cell level |
| CN111378576A (en) * | 2018-12-28 | 2020-07-07 | 上海大学 | Intestinal tract simulation chip and application thereof |
| CN111378576B (en) * | 2018-12-28 | 2023-10-31 | 上海大学 | Intestinal tract simulation chip and application thereof |
| CN113101992A (en) * | 2021-05-08 | 2021-07-13 | 苏州珀罗汀生物技术有限公司 | Protein synthesis screening method |
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Inventor after: Gan Mingzhe Inventor after: Wang Dandan Inventor after: Cui Jinhui Inventor before: Gan Mingzhe Inventor before: Wang Dandan Inventor before: Luo Dan |