CN109097276A - Non-contact cell co-culture device, manufacturing method and cell culture method - Google Patents
Non-contact cell co-culture device, manufacturing method and cell culture method Download PDFInfo
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- CN109097276A CN109097276A CN201810941528.7A CN201810941528A CN109097276A CN 109097276 A CN109097276 A CN 109097276A CN 201810941528 A CN201810941528 A CN 201810941528A CN 109097276 A CN109097276 A CN 109097276A
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- 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
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B33—ADDITIVE MANUFACTURING TECHNOLOGY
- B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
- B33Y30/00—Apparatus for additive manufacturing; Details thereof or accessories therefor
Abstract
The invention discloses a non-contact cell co-culture device, a manufacturing method and a cell culture method. The non-contact cell co-culture device comprises a PDMS culture chamber, wherein two or more culture channels are formed in the PDMS culture chamber; each culture channel comprises a horizontal groove and two vertical channels; a mold is prepared by a 3D printing technology; then PDMS is poured into the mold to manufacture the PDMS culture chamber; when cells are cultured, the PDMS culture chamber is arranged on a substrate; various co-cultured cell suspensions are injected into the culture channels; after the cells in the cell suspensions are adhered, the PDMS culture chamber is removed, and culture is continued to form a co-culture system of various cells. According to the non-contact cell co-culture device, the manufacturing method and the cell culture method, two or more cell co-culture systems can be conveniently established, and the mutual effect between different cells is favorably researched; the cell co-culture system established by the method is high in production efficiency, economic, convenient, simple in process and easy for large-scale production.
Description
Technical field
The invention belongs to Biotechnology field more particularly to a kind of contactless cell co-cultivations based on 3D printing
Device.
Background technique
The organic whole that organism is made of various kinds of cell, by influencing each other between different cells, Coordinated Play is made
With, maintain organ, tissue it is normal, influencing each other between cell has been a hot spot of research.There are phases between different cells
Mutually influence, such as: cell communication between oligodendroglia and neuronal cell influences the formation of myelin, cardiac muscle cell at
Influencing each other between fibrocyte participates in the growth course of heart, the interaction between Dendritic Cells and T lymphocyte
Participate in the antigen deduction process, etc. in immune response.There is also influencing each other between allogenic cell different conditions, such as.Depending on
Apoptotic signal in nethike embrane layer can be spread within the organization, make the normal cell around apoptotic cell that apoptosis occur.Myocardial infarction
In the process, dead signal can be connected by gap and be transmitted, and make the normal cell around dead myocardial cells that apoptosis or bad occur
It waits indefinitely.Influencing each other extensively between cell, plays a significant role under organism physiology and pathological state, is worth carry out system
Research.
Research cell and intercellular interaction can be realized by the co-cultivation of different types of cell.Currently,
The method that cell co-cultures mainly has direct contact type co-cultivation and non-direct contact type to co-culture.Direct contact type co-cultures
Different types of cell is mixed to carry out co-incubation in the same cultivating system, contacts two kinds of cells directly,
And by paracrine, Autocrine generates Porcine HGF etc. and interacts.The disadvantage is that different types of cell
Occur directly to contact, cannot thoroughly separate, is unfavorable for subsequent analysis observation.Non-direct contact type co-cultivation is by two kinds or two kinds
Above cell is inoculated into respectively on different carrier or position, and then these cells are placed in the same culture environment, altogether
Influence of the one to another person is interacted by the cell factor of paracrine in culture environment, and two kinds of cells do not contact with each other,
I.e. a kind of cell factor of cell release can reach another cell growth position by diffusion and have an impact to it, because
This, will not influence subsequent detection convenient for observation.But non-direct contact type co-culture model preparation process is complicated, operation compared with
It is cumbersome.Therefore, there is presently no a kind of easily and economically material benefit and convenient for the co-cultivation of two or more cells of large scale preparation
The model of system.
Summary of the invention
Goal of the invention: in order to overcome the deficiencies in the prior art, the present invention provides a kind of non-based on 3D printing and connects
Touch cell co-cultures device and manufacturing method and cell culture processes.Device is co-cultured using the cell, can guarantee different cells
Between non-direct contact, while may be implemented to study and interact between different cells.
Technical solution: to achieve the above object, the technical solution adopted by the present invention are as follows:
A kind of contactless cell co-cultivation device, including PDMS culturing room, the PDMS culturing room include culture chamber body, training
It supports and offers more than two culture channels in chamber body, the culture channel includes horizontal groove and two Vertical Channels, and two
Vertical Channel is respectively Vertical Channel one, Vertical Channel two, and one end of Vertical Channel one connects with one end of horizontal groove, another
End setting forms adding mouth in PDMS culture chamber surface.One end of Vertical Channel two connects with the other end of horizontal groove, vertically
The setting of two other end of channel forms outlet in PDMS culture chamber surface.The distance between two adjacent horizontal grooves are 90-
110 μm, the height of horizontal groove is 290-310 μm.
Further: further including substrate, the PDMS culturing room is arranged on substrate, so that Vertical Channel one, level are recessed
Slot, Vertical Channel two are in contact with substrate and penetrate through culture channel.
It is preferred: to be parallel to each other between the horizontal groove.
Preferred: the length between two adjacent horizontal grooves is inconsistent.
Preferred: the shape of the horizontal groove and Vertical Channel is cuboid.
A method of contactless cell is manufactured based on 3D printing technique and co-cultures device, comprising the following steps:
Step 1, mold is prepared by 3D printer printing by polylactic acid PLA, the mold includes more than two cultures
Channel forming mould and mold base (4), the culture channel shaping mould are arranged on mold base (4), the mold base (4)
Surrounding be provided with coaming plate.The culture channel shaping mould includes two vertical columns and horizontal projection item, two vertical columns
It is separately positioned on the both ends of horizontal projection item, wherein the distance between two adjacent horizontal projection items are 90-110 μm, horizontal
The height of raised item is 290-310 μm.
Step 2, polydimethylsiloxane monomer and its crosslinking agent are sufficiently mixed according to the volume ratio of 10:1, degasification
It is cast on mold after bubble, by heating 1.8-2.2h molding at 75-85 DEG C, then mold is extracted, obtains PDMS culture
Room.
Step 3, hydrophilic surface modification is carried out to PDMS culturing room using reactive ion etching machine, obtains modified PDMS
Culturing room.
Preferred: when 3D printing prepares mold in step 1,3D printing model is by Solidworks software building, by STL
Input 3D printer forms printable order after format conversion.
Further: PDMS culturing room obtained in step 2 being put into chloroform chemical solvent and polishes 10-30s, takes out and uses
Water rinses and dries, the PDMS culturing room after being polished.
Further: the modified PDMS culturing room of step 3 being transferred on the glass slide for being placed with substrate, PDMS culture
It is in close contact between room and substrate.
A kind of cell culture processes co-culturing device using contactless cell, comprising the following steps:
Step A prepares the various cell suspensions co-cultured, wherein the type co-cultured is not more than the quantity of culture channel.
PDMS culturing room is transferred on the glass slide for being placed with substrate by step B, close between PDMS culturing room and substrate
Contact, so that Vertical Channel one, horizontal groove, Vertical Channel two are in contact with substrate and penetrate through culture channel.
A kind of cell suspension is injected into the culture channel after a perforation by step C, after having injected cell suspension, is protected
Without the culture channel for being not injected into cell suspension between the culture channel of card injection cell suspension.
The PDMS culturing room for being filled with cell suspension and glass slide are placed in incubator culture together and carry out by step D
Culture, moves back except PDMS culturing room to the cell adhesion in cell suspension and continues to cultivate, form the total training of various cells
The system of supporting.
The present invention compared with prior art, has the advantages that
(1) present invention can easily establish the contactless co-culture system of two or more cells very much.
(2) cell culture fluid can be continuously added into the channel of co-culture system by the present invention, realize the training altogether of cell dynamic
It supports.
(3) present invention co-cultures mold and utilizes 3D printing technique, passes through Solidworks software modeling and is converted to STL and beats
Print can execute order, and the rapid shaping of mold, simple process, high production efficiency may be implemented.
(4) culturing room that the present invention is co-cultured using biocompatible material PDMS as cell, material, which obtains, to be easy, and is kept away
Exempt from using expensive material.
(5) the present invention is based on the molds of 3D printing convenient for removing, and can reuse.
(6) the present invention is based on the cell of 3D printing co-culture device it is easy to operate, be conducive to be mass produced.
Detailed description of the invention
Fig. 1 is that contactless cell of the invention co-cultures device and removes the mold structure diagram of coaming plate.
Fig. 2 is the left view and C-C cross-sectional view that contactless cell co-cultures device in Fig. 1, wherein Fig. 2 a is contactless
Cell co-cultures the left view of device, and Fig. 2 b is the C-C cross-sectional view of Fig. 2 a.
Fig. 3 is the left view and B-B cross-sectional view that the mold of coaming plate is removed in Fig. 1, wherein Fig. 3 a is the mold for removing coaming plate
Left view, Fig. 3 b be Fig. 3 a B-B cross-sectional view.
Fig. 4 is the left perspective view and F-F cross-sectional view, D-D cross-sectional view that contactless cell co-cultures device in Fig. 1, wherein figure
4a is the left perspective view that contactless cell co-cultures device, and Fig. 4 b is the F-F cross-sectional view of Fig. 4 a, and Fig. 4 b is the D-D section view of Fig. 4 c
Figure.
Fig. 5 is the left perspective view and G-G cross-sectional view, E-E cross-sectional view that the mold of coaming plate is removed in Fig. 1, wherein Fig. 5 a is non-
Contact type cell co-cultures the left perspective view of device, and Fig. 5 b is the G-G cross-sectional view of Fig. 5 a, and Fig. 5 b is the E-E cross-sectional view of Fig. 5 c.
Fig. 6 is that the contactless cell of embodiment 1 co-cultures device and removes the mold structure diagram of coaming plate.
Fig. 7 is that the contactless cell of embodiment 2 co-cultures device and removes the mold structure diagram of coaming plate.
In above-mentioned figure: 1- horizontal projection strip, 2- horizontal projection billet, the vertical column of 3-, 4- mold base, 5- lead to vertically
Road one, the horizontal long recess of 6-, the horizontal short groove of 7-, 8- cultivate chamber body, and 9- many cells co-culture first passage, and 10- many cells are total
Second channel is cultivated, 11- many cells co-culture third channel, and 12 be PDMS culturing room, and 13 be mold.
Specific embodiment
In the following with reference to the drawings and specific embodiments, the present invention is furture elucidated, it should be understood that these examples are merely to illustrate this
It invents rather than limits the scope of the invention, after the present invention has been read, those skilled in the art are to of the invention various
The modification of equivalent form falls within the application range as defined in the appended claims.
A kind of contactless cell co-cultivation device, as shown in Fig. 1,2,4, including PDMS culturing room 12, PDMS culturing room 12
For cultivating different cells, the PDMS culturing room 12 includes culture chamber body 8, cultivates and offers more than two trainings in chamber body 8
Support channel, culture channel quantity can according to need flexible and changeable, and the culture channel includes that horizontal groove and two are vertical logical
Road, two Vertical Channels are respectively Vertical Channel one, Vertical Channel two, one end of Vertical Channel one and one end phase of horizontal groove
It connects, other end setting forms adding mouth in PDMS culture chamber surface.The other end phase of one end of Vertical Channel two and horizontal groove
It connects, the setting of two other end of Vertical Channel forms outlet in PDMS culture chamber surface, and cell suspension is added by adding mouth, passes through
Vertical Channel one, horizontal groove, Vertical Channel two, cell suspension is flowed out from outlet, therefore dynamic cell culture may be implemented.
The distance between two adjacent horizontal grooves are 90-110 μm, and the height of horizontal groove is 290-310 μm.
It further include substrate, substrate is sheet glass, and the PDMS culturing room is arranged on substrate, so that Vertical Channel one, water
Flat grooves, Vertical Channel two are in contact with substrate and penetrate through culture channel, channel that culture channel is conveyed as cell suspension and
Culture channel.
It is parallel to each other between the horizontal groove.Length between two adjacent horizontal grooves is inconsistent.It is divided into level
Long recess 6, horizontal short groove 7.The shape of the horizontal groove and Vertical Channel is cuboid.
PDMS culturing room of the invention uses biocompatible material PDMS.This cost of material is low, using simple, and has
A kind of the features such as good chemical inertness, it has also become polymer material for being widely used in micro-fluidic equal fields.PDMS is poured on
On mold and heating solidifies PDMS.When mold is removed, the channel design of mold will be transferred on PDMS.Utilize reaction
Ion etching machine carries out hydrophilic surface modification, obtains modified PDMS culturing room.It can carry out cell co-cultivation.
Mutually control, each cultivating system have a plurality of horizontal culture channel, can be divided into different groups the present invention for convenience.
Horizontal groove (horizontal channel) length in different groups is different, to be separately added into cell sample.Interchannel is mutually not through respectively
Cultivate different cells.It can use different PDMS culturing room when experiment, carry out two or more cell co-cultivations, respectively to study difference
It is anisotropic.
In each cultivating system, PDMS culturing room has a plurality of horizontal culture channel, and is parallel to each other.Channel and channel it
Between only at a distance of 100 μm, 300 μm of channel height.Channel generally rectangular parallelepiped structure.
A method of contactless cell is manufactured based on 3D printing technique and co-cultures device, comprising the following steps:
Step 1, mold being prepared by 3D printer printing by polylactic acid PLA, mold makes PDMS culturing room for demoulding,
When 3D printing prepares mold, 3D printing model inputs 3D printer after the conversion of STL format by Solidworks software building
Form printable order.The mold includes more than two culture channel shaping moulds and mold base 4, the culture channel at
Pattern is arranged on mold base 4, and the surrounding of the mold base 4 is provided with coaming plate.The culture channel shaping mould includes two
A vertical column 3 and horizontal projection item, two vertical columns 3 are separately positioned on the both ends of horizontal projection item, wherein adjacent two
The distance between a horizontal projection item is 90-110 μm, and the height of horizontal projection item is 290-310 μm.PLA is molten in printing
Melt state, is formed after cooling.Length between two adjacent horizontal projection items is inconsistent.It is divided into horizontal projection strip 1, it is horizontal
Raised billet 2.Mold may be reused.
Step 2, polydimethylsiloxane monomer and its crosslinking agent are sufficiently mixed according to the volume ratio of 10:1, degasification
It is cast on mold after bubble, by heating 1.8-2.2h molding at 75-85 DEG C, then mold is extracted, obtains PDMS culture
Room.Obtained PDMS culturing room is put into chloroform chemical solvent and polishes 10-30s, taking-up is rinsed with water and dries, and is polished
PDMS culturing room afterwards.
Step 3, hydrophilic surface modification is carried out to PDMS culturing room using reactive ion etching machine, obtains modified PDMS
Culturing room.Modified PDMS culturing room is transferred on the glass slide for being placed with substrate, and keeps PDMS culturing room and substrate
Between be in close contact.PDMS culturing room is made of PDMS culture channel, and PDMS culturing room may be reused.
A kind of cell culture processes co-culturing device using contactless cell, comprising the following steps:
Step A prepares the various cell suspensions co-cultured, wherein the type co-cultured is not more than the quantity of culture channel.
PDMS culturing room is transferred on the glass slide for being placed with substrate by step B, and keep PDMS culturing room and substrate it
Between be in close contact so that Vertical Channel one, horizontal groove, Vertical Channel two are in contact with substrate and penetrate through culture channel.
A kind of cell suspension is injected into the culture channel after a perforation by step C, after having injected cell suspension, is protected
Without the culture channel for being not injected into cell suspension between the culture channel of card injection cell suspension.
The PDMS culturing room for being filled with cell suspension and glass slide are placed in incubator culture together and carry out by step D
Culture, moves back except PDMS culturing room to the cell adhesion in cell suspension and continues to cultivate, since various cells are being cultivated
It is cultivated in channel, therefore, (removes culture channel) after removing PDMS culturing room, whithin a period of time (3-15 days), respectively
Kind cell also maintains former culture channel shape to be cultivated, due to there is gap between culture channel, after removing PDMS culturing room, respectively
Kind cell does not also contact with each other, and cell factor is diffused by cell culture fluid, forms the co-culture system of various cells.
PDMS culturing room, convenient for removing, does not influence the cell co-cultured in substrate during cell co-cultures after removal
Growth.PDMS culturing room can be very easy building cell co-culture system, it is easily operated and large-scale production.
Embodiment 1
The present embodiment is co-cultured using two kinds of cells:
(1) 3D printing mold process: setting 3D printer software parameter.Thickness: 0.1mm；Wall thickness: 0.8mm；Bottom/top
Thickness degree 0.8mm；Packed density: 100%；Print speed: 30mm/s；Print temperature: 200 degrees Celsius；Material diameter:
1.75mm；Flow of material: 100%.Mould structure is designed by CAD software Solid Works, and is converted to STL formatted file (figure
1-3), then by 3D printer it prints, printed material used is PLA.
Polishing process after printing: after the completion of printing, there are part flaws for die surface, first can gently be wiped with pocket knife, then
Mold is put into chloroform chemical solvent and polishes 10-30s.Mold is taken out, is rinsed and is air-dried with tap water, is i.e. acquisition surface is smooth
Seamless PLA mold.
(2) PDMS culturing room forming process: PDMS monomer and its crosslinking agent are sufficiently mixed according to the volume ratio of 10:1, removed
It is cast on mold after bubble, 2h is heated at 80 DEG C, PDMS is removed from mold then and obtains bottom with horizontal groove
PDMS culturing room.The PDMS culturing room of preparation will guarantee that bottom is smooth, without obvious gully.It is transferred on glass slide after molding,
Guarantee PDMS culturing room bottom on glass slide glass completely it is seamless agree with, not will cause hydraulic fluid side leakage or leakage.
(3) two kinds of cells co-culture: duct (shown in Fig. 6) is separately added into Shi Wanxi with syringe at the top of from PDMS culturing room
Born of the same parents' suspension and Dorsal Root Ganglion Neurons suspension are moved back to cell adhesion except co-cultivation mold after being put into incubator culture 4h, and after
It is continuous to be cultivated, form the co-culture system of two kinds of cells of schwann cells and Dorsal Root Ganglion Neurons.Incubation must guarantee carefully
Born of the same parents' suspension is retained in respective channel, and cell is without internal side leakage and outside side leakage phenomenon.
Embodiment 2
The present embodiment is co-cultured using three kinds of cells,；
(1) 3D printing mold process: co-culturing three kinds of cells, only need to than the two kinds more one group of channel of cells co-cultivation.
For convenience of differentiation, three kinds of cell cultivation level passage lengths are different, and any one horizontal channel two sides are horizontal length
Different channels.3D printer software parameter is set.Thickness: 0.2mm；Wall thickness: 1.0mm；Bottom/top layers thickness 1mm；It fills close
Degree: 100%；Print speed: 30mm/s；Print temperature: 200 degrees Celsius；Material diameter: 1.75mm；Flow of material: 100%.By
CAD software Solid Works designs mould structure, and is converted to STL formatted file (Fig. 4), then printed by 3D printer, used
Printed material is PLA.
Polishing process after printing: after the completion of printing, there are part flaws for die surface, first can gently be wiped with pocket knife, then
Mold is put into chloroform chemical solvent and polishes 10-30s.Mold is taken out, is rinsed and is air-dried with tap water, is i.e. acquisition surface is smooth
Seamless PLA mold.
(2) PDMS culturing room forming process: PDMS monomer and its crosslinking agent are sufficiently mixed according to the volume ratio of 10:1, removed
It is cast on mold after bubble, 2h is heated at 80 DEG C, PDMS is removed from mold then and obtains bottom with channel
PDMS culturing room.The DMS culturing room of preparation will guarantee that bottom is smooth, without obvious gully.It is transferred on glass slide, to protect after molding
Card PDMS culturing room bottom on glass slide glass completely it is seamless agree with, not will cause hydraulic fluid side leakage or leak outside.
(3) three kinds of cells co-culture: duct is (as shown in fig. 7, many cells co-culture first passage at the top of from PDMS culturing room
9, many cells co-culture second channel 10, and many cells co-culture third channel 11) it is separately added into endothelial cell suspension with syringe,
Smooth muscle cell suspension and fibroblast suspension are put into incubator culture 4h and wait for that cell adhesion is moved back except DMS culturing room, and after
It is continuous to be cultivated, form the co-culture system of three kinds of endothelial cell, smooth muscle cell and fibroblast cells.Incubation palpus
Guarantee that cell suspension is retained in respective channel, cell is without internal side leakage and outside side leakage phenomenon.
Mold of the invention is prepared by polylactic acid (PLA) by 3D printing, has certain intensity, and will not train to PDMS
Feeding room has an impact.PDMS culturing room is made using dimethyl silicone polymer (PDMS) material, by be cast to after mold again into
It is formed in row demoulding to glass slide.After cast molding, mold is extracted.It include multiple sizes above PDMS culturing room after molding
The injection port and outlet of consistent cell suspension are rectangular recess below PDMS culturing room, above cell suspension sample introduction
Mouth and outlet are with lower recess connection.The quantity of injection port and outlet and groove is not fixed (culture channel), can be with
It is flexible and changeable, thus co-culture cell category be also possible to it is a variety of.Different sample holes can be passed through different types of cell.By
Injection port opens below into cell suspension, can remove PDMS culturing room after cell culture for a period of time, form two or more
The co-cultivation of cell.The present invention can easily establish two or more cell co-culture systems, and it is different thin to be conducive to research
Interaction between born of the same parents, the cell co-culture system high production efficiency that this method is established is economical convenient, and simple process, easily
Large-scale production.
The above is only a preferred embodiment of the present invention, it should be pointed out that: for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (10)
1. a kind of contactless cell co-cultures device, it is characterised in that: including PDMS culturing room, opened up in the PDMS culturing room
There are two above culture channel, the culture channel includes horizontal groove and two Vertical Channels, two Vertical Channel difference
For Vertical Channel one, Vertical Channel two, one end of Vertical Channel one connects with one end of horizontal groove, and the other end is arranged in PDMS
It cultivates chamber surface and forms adding mouth；One end of Vertical Channel two connects with the other end of horizontal groove, two other end of Vertical Channel
Setting forms outlet in PDMS culture chamber surface；The distance between two adjacent horizontal grooves are 90-110 μm, horizontal recessed
The height of slot is 290-310 μm.
2. contactless cell co-cultures device according to claim 1, it is characterised in that: it further include substrate, the PDMS training
It supports room to be arranged on substrate, so that Vertical Channel one, horizontal groove, Vertical Channel two are in contact with substrate and pass through culture channel
It is logical.
3. contactless cell co-cultures device according to claim 2, it is characterised in that: mutually flat between the horizontal groove
Row.
4. contactless cell co-cultures device according to claim 2, it is characterised in that: between two adjacent horizontal grooves
Length it is inconsistent.
5. contactless cell co-cultures device according to claim 2, it is characterised in that: the horizontal groove and Vertical Channel
Shape be cuboid.
6. a kind of method for co-culturing device based on contactless cell described in 3D printing technique manufacturing claims 1, feature
It is: the following steps are included:
Step 1, mold is prepared by 3D printer printing by polylactic acid PLA, the mold includes more than two culture channels
Shaping mould and mold base (4), the culture channel shaping mould are arranged on mold base (4), and the four of the mold base (4)
Week is provided with coaming plate；The culture channel shaping mould includes two vertical columns and horizontal projection item, two vertical column difference
The both ends of horizontal projection item are set, wherein the distance between two adjacent horizontal projection items are 90-110 μm, horizontal projection
The height of item is 290-310 μm；
Step 2, it will be cast on mold after polydimethylsiloxane monomer and its crosslinking agent mixing bubble removing, in 75-85
By heating 1.8-2.2h molding at DEG C, then mold is extracted, obtains PDMS culturing room；
Step 3, hydrophilic surface modification is carried out to PDMS culturing room using reactive ion etching machine, obtains modified PDMS culture
Room.
7. method according to claim 6, it is characterised in that: when 3D printing prepares mold in step 1,3D printing model by
Solidworks software building, input 3D printer forms printable order after the conversion of STL format.
8. method according to claim 7, it is characterised in that: PDMS culturing room obtained in step 2 is put into chloroform chemistry
10-30s is polished in solvent, taking-up is rinsed with water and dries, the PDMS culturing room after being polished.
9. method according to claim 6, it is characterised in that: be transferred to the modified PDMS culturing room of step 3 and be placed with
On the glass slide of substrate, make to be in close contact between PDMS culturing room and substrate.
10. a kind of cell culture processes for co-culturing device using contactless cell described in claim 1, which is characterized in that
The following steps are included:
Step A prepares the various cell suspensions co-cultured, wherein the type co-cultured is not more than the quantity of culture channel；
PDMS culturing room is transferred on the glass slide for being placed with substrate by step B, so that Vertical Channel one, horizontal groove, vertical
Channel two is in contact with substrate penetrates through culture channel；
A kind of cell suspension is injected into the culture channel after a perforation by step C, after having injected cell suspension, guarantees note
Enter between the culture channel of cell suspension without the culture channel for being not injected into cell suspension；
The PDMS culturing room for being filled with cell suspension and glass slide are placed in incubator culture together and cultivate by step D,
It moves back to the cell adhesion in cell suspension except PDMS culturing room and continues to cultivate, form the co-cultivation body of various cells
System.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111621468A (en) * | 2020-05-21 | 2020-09-04 | 上海慧灯医疗科技有限公司 | Method for preparing induced pluripotent stem cell-derived cardiomyocytes and fixing method and application thereof |
| CN113512497A (en) * | 2021-08-12 | 2021-10-19 | 浙江大学 | 3D printing culture tank, culture system and detection system for in-vitro intestinal living body culture and application |
| CN114107047A (en) * | 2021-11-18 | 2022-03-01 | 北京理工大学 | Multi-cell co-culture three-dimensional micro-scaffold embedded flow channel based on two-photon lithography |
| CN114752494A (en) * | 2022-03-24 | 2022-07-15 | 四川大学 | Cell culture chamber, adjustable cell mechanical stimulation culture device and manufacturing method thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN203923211U (en) * | 2014-06-24 | 2014-11-05 | 谢一泓 | A kind of co-culture of cells orifice plate |
| CN106520698A (en) * | 2016-01-05 | 2017-03-22 | 苏州汶颢芯片科技有限公司 | Cell non-contact culture method based on micro-fluidic chip |
| CN107267388A (en) * | 2017-08-08 | 2017-10-20 | 清华大学 | A kind of multi-functional cell co-cultures capsule and its application |
-
2018
- 2018-08-17 CN CN201810941528.7A patent/CN109097276B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN203923211U (en) * | 2014-06-24 | 2014-11-05 | 谢一泓 | A kind of co-culture of cells orifice plate |
| CN106520698A (en) * | 2016-01-05 | 2017-03-22 | 苏州汶颢芯片科技有限公司 | Cell non-contact culture method based on micro-fluidic chip |
| CN107267388A (en) * | 2017-08-08 | 2017-10-20 | 清华大学 | A kind of multi-functional cell co-cultures capsule and its application |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111621468A (en) * | 2020-05-21 | 2020-09-04 | 上海慧灯医疗科技有限公司 | Method for preparing induced pluripotent stem cell-derived cardiomyocytes and fixing method and application thereof |
| CN113512497A (en) * | 2021-08-12 | 2021-10-19 | 浙江大学 | 3D printing culture tank, culture system and detection system for in-vitro intestinal living body culture and application |
| CN114107047A (en) * | 2021-11-18 | 2022-03-01 | 北京理工大学 | Multi-cell co-culture three-dimensional micro-scaffold embedded flow channel based on two-photon lithography |
| CN114107047B (en) * | 2021-11-18 | 2023-06-23 | 北京理工大学 | Multicellular co-culture three-dimensional micro-stent embedded runner based on two-photon lithography |
| CN114752494A (en) * | 2022-03-24 | 2022-07-15 | 四川大学 | Cell culture chamber, adjustable cell mechanical stimulation culture device and manufacturing method thereof |
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