CN207760352U

CN207760352U - Media to cell culture modules, culture solution module, cell cultivation chip and cell culture cabinet

Info

Publication number: CN207760352U
Application number: CN201721772971.3U
Authority: CN
Inventors: 钱其军; 周进; 师传胤; 颜开冬
Original assignee: Shanghai Cell Therapy Engineering Technology Research Center Group Co Ltd; Shanghai Cell Therapy Research Institute; Shanghai Baize Medical Instrument Co Ltd
Current assignee: Shanghai Baize Medical Instrument Co ltd; Shanghai Cell Therapy Research Institute; Shanghai Cell Therapy Group Co Ltd
Priority date: 2017-12-18
Filing date: 2017-12-18
Publication date: 2018-08-24
Anticipated expiration: 2027-12-18
Also published as: WO2019120136A1

Abstract

The utility model is related to media to cell culture modules, culture solution module, cell cultivation chip and cell culture cabinet, the media to cell culture modules includes：The closed ontology being in communication with the outside only is exported by culture solution import, culture solution outlet, cell liquid import and cell liquid；Be located within the body the culture sap cavity and lumen in portion, wherein the culture sap cavity include it is independent respectively with the culture sap cavity of culture solution inlet communication and with the culture sap cavity of culture solution outlet, the lumen and the cell liquid import and cell liquid outlet；Sealing block, for being separated to form the lumen and culture sap cavity；Be placed in it is in lumen, on wall have do not allow cell by micropore hollow fibre filament；Wherein, the hollow fibre filament passes through the sealing block, and the both ends of hollow fibre filament are connected to the culture sap cavity of the culture sap cavity of culture solution input end and culture solution outlet end respectively.

Description

Media to cell culture modules, culture solution module, cell cultivation chip and cell culture cabinet

Technical field

The utility model belongs to biomedical field of cell culture, and in particular to media to cell culture modules, culture solution module, cell Cultivate chip and cell culture cabinet.

Background technology

Nowadays, biological immune treatment has become a kind of emerging means for capturing tumor and cancer, is biological therapy technical field A key technology, have well research and development and application prospect.Cell therapy in biological immune treatment is from patient's body Extraction sub-elects a certain amount of immunocyte, and in carrying out amplification cultivation in vitro, which may include specific stimulating factor addition And/or the use of transgenic technology, after cell is expanded to certain scale or quantity, according to the course for the treatment of of patient needs feed back into Itself is in vivo.During the entire course for the treatment of, the amplification in vitro culture of immunocyte is the important component that technology is realized, rationally Cell culture apparatus be to realize the key factor that safely, effectively expands of immunocyte.Existing cell therapy mechanism is for cell In vitro culture mostly uses common porous plate, Tissue Culture Flask or permeability type culture bag and carries out static culture as container, this Mode can only provide a static growth environment to cell, and cells grown environment and internal true environment difference are larger, and Culture medium is only carried out in incubation to add, is not replaced, and the product of cells grown metabolism easily accumulates, to influence immunocyte Growth rate, form size, cell phenotype, expression status and cell quantity and vigor etc., easily limitation and reduce immune The effect for the treatment of；And artificial container of repeatedly opening is needed to be added culture solution and the operation of other confactors in production, increase The risk for having added sample to pollute also proposes the technology for cultivating operating personnel and requires higher.Meanwhile more people Work operation link is also easy the consistency of quality between influence immunocyte culture products batch.

Immunocyte can be divided into attached cell and suspension cell according to the growth pattern of cell, and attached cell depends on culture It is grown on wall of a container, suspension cell suspension growth in the culture solution of culture vessel.Ideal cell culture apparatus needs energy Enough while this two classes cell is taken into account, to reduce the input for instrument or consumptive material, raising utilization rate of equipment and installations.Immunocyte is in body In interior growth course, the blood of flowing can not only feed vegetative cell, can also take away the waste that cell secretion generates, but these The dynamic environment such as the transmission of nutriment, the discharge for secreting waste are difficult to simulate in common static culture apparatus.Exist at present Have some in field of cell culture and can be used for devices and technology of dynamic cell culture, disclosure satisfy that cells grown space, The functions such as liquid, such as rolling bottle, bioreactor (bioreactor), hollow fiber conduit are changed, but the design of these equipment is complicated, operation Cumbersome, wherein rolling bottle is simply possible to use in attached cell culture, and culture space is limited, more and more thin by extensive multilayer at present Born of the same parents factory is replaced；Culture tank can meet the cell culture of large volume, but traditional stainless steel or glass tank body produces every time Be required for being cleaned and being sterilized before, can not suitable for cell therapy clinical application individuation cell culture.Other classes The reactor that the reactor of type such as wave is waved uses medical disposable material, but it is multi-party to be limited by equipment instrument, toxigenic capacity etc. Face factor, therefore be not widely accepted.Existing doughnut training method often exclusively carries out design for certain cell, Application range is narrow.

And the culture for developing into cell of microfluidic chip technology provides new platform, micro-fluidic chip has space body Product is small, integrated level is high, the advantages such as disposable.But there are still culture solutions for current existing related micro fluidic chip developmental achievement The problems such as inconvenient, cell culture area is endless totally-enclosed, gas exchanges are incomplete is replaced, and is intended for the survey of few cells mostly The real-time dynamic monitoring direction of examination culture and cell state, is not particularly suited for the large-scale production of cell.

CN158703A discloses a kind of minicell hollow fiber reactor, which is that the pre-buried filling one of cell is small In section doughnut filament, then it is placed in the closed ventilative silicone tube for containing culture solution.The device is by means of hollow fibre The transmission that silk micropore carries out nutriment is tieed up, Related Drug can be can be applied to by the culture of filament inner cell to certain scale The research of object.The device is the pharmacological research applied to few cells, for large-scale clinical cytology culture be difficult to meet with It is required that.

CN103667054A discloses a kind of integrated micro-flow control cell cultivation chip, and the chip is by chip body, with phase The accessories such as temperature control component, Micropump, micro-valve and the control circuit of pass are integrated in one.Chip body is bonded by multilayer, including Cell culture area, culture solution memory block, waste etc., are connected between different zones by microfluidic circuit.The apparatus integration Integrated, fluid channel inner cavity is closed, is suitable for the culture studies of few cells, but flow path designs are simple, for immunocyte Culture environment simulation is not comprehensive enough, and ignores the gas exchanges condition of cell culture, and it is thin to be not directly applicable clinical immunization The culture of born of the same parents.

CN105754856A discloses a kind of drawer type three-dimensional class blood vessel tissue culture organ chip, which is by taking out Drawer casket, sealing plate, fixed frame, class vascularization three-dimensional tissue and its relevant inlet and outlet pipe fitting are constituted.The level of integrated system is high, can Dismounting or sealing, automation monitoring in real time, simulate emphatically the stabilization adhere-wall culture of class blood vessel three-dimensional tissue, and use transparent air-permeable Formula chip material is conducive to the growth and monitoring in real time of relevant cell, but the chip is only the training for being intended for blood vessel tissue It supports, passability is not strong.

Invention content

The utility model is directed to the shortcoming of existing various types of cells cultural method and device, in conjunction with doughnut and culture The advantage of chip, provide it is a kind of it is simple in structure, operation is convenient, the media to cell culture modules suitable for immunocyte and its culture dress Set, have spatial volume is small, culture density is high, can automation control, the closing of chip culture space, chip assembly is easily assembled more It changes, the features such as bio-safety performance height.The cell culture apparatus is exempted from by cultivating in the abundant simulated blood vessel of built-in chip type filament The growing environment of epidemic disease cell establishes fairly large 3D cell culture, and panimmunity is thin suitable for cell therapy clinical application The culture of born of the same parents, such as the DC cells (Dendritic Cells) of adherent type, a variety of T lymphocytes (CIK, CTL, the TIL of suspension type Deng) culture.

The media to cell culture modules of the utility model includes：

It is only in communication with the outside by culture solution import, culture solution outlet, cell liquid import and cell liquid outlet closed Ontology；

Be located within the body the culture sap cavity and lumen in portion, wherein the culture sap cavity include it is independent respectively with culture The culture sap cavity of liquid inlet communication and the culture sap cavity with culture solution outlet, the lumen and the cell liquid import and Cell liquid outlet；

Sealing block, for being separated to form the lumen and culture sap cavity；With

Be placed in it is in lumen, on wall have do not allow cell by micropore hollow fibre filament；Wherein, described hollow Filament passes through the sealing block, and the both ends of hollow fibre filament go out with the culture sap cavity and culture solution of culture solution input end respectively The culture sap cavity at mouth end is connected to.

In certain embodiments, the volume of the culture sap cavity is 0.2~0.5L, preferably 0.3~0.35L.

In certain embodiments, the volume of the lumen is 0.4~1L, preferably 0.6~0.8L.

In certain embodiments, the length of the hollow fibre filament is 130~170mm, 20~50 μm of thickness of pipe wall, tube wall The aperture of upper micropore is less than 1 μm.

In certain embodiments, the ontology and sealing block are transparent biocompatible materials.

In certain embodiments, the culture sap cavity is located at lumen both sides, is separated by sealing block, hollow fibre filament Across lumen to be connected to the culture sap cavity of lumen both sides.

In certain embodiments, the culture sap cavity is all located at lumen the same side, between two culture sap cavities and It is separated by sealing block between culture sap cavity and lumen.

In certain embodiments, culture solution import and culture solution outlet inside are respectively equipped with sedimeter, to Broth filtrate, the pollution that may be introduced when culture solution inlet and outlet being avoided to be connected to culture solution module.

In certain embodiments, culture solution import, culture solution outlet are all made of medical rubber stopper and are sealed.

In certain embodiments, the culture sap cavity with culture solution inlet communication has culture sap cavity import, the training The import of nutrient solution chamber and the culture solution inlet communication；There is the culture sap cavity with culture solution outlet culture sap cavity to go out Mouthful, the culture sap cavity outlet and the culture solution outlet.

In certain embodiments, the culture sap cavity import is located in the ontology, passes through intrinsic processing flow path With the culture solution inlet communication.

In certain embodiments, the culture sap cavity outlet is located in the ontology, and the media to cell culture modules also wraps The processing flow path for being located at the body interior and the optional culture fluid path transfer tube pipeline for being located at the body exterior are included, it is described Sap cavity outlet is cultivated by the processing flow path and the optional culture fluid path transfer tube pipeline to connect with culture solution outlet It is logical.

In certain embodiments, there is the lumen lumen import and lumen to export, respectively with cell liquid into Mouth and cell liquid outlet.

In certain embodiments, the lumen import is each provided at the body interior with lumen outlet, described thin It further includes being located at the cell fluid path transfer tube pipeline of the body exterior that born of the same parents, which cultivate module, wherein the lumen import with it is thin Cell outlet is described thin respectively by being located at processing flow path and the cell liquid import and cell liquid outlet of the body interior Pass through the cell fluid path transfer tube pipeline connection between cytosol import and cell liquid outlet.

In certain embodiments, the media to cell culture modules has culture solution flow path and cell liquid flow path；Wherein, it cultivates Liquid stream road includes culture solution import, processing flow path successively, cultivates sap cavity import, in the culture sap cavity of input end, hollow fibre filament Portion, the culture sap cavity of outlet end, the outlet of culture sap cavity, processing flow path, optional culture fluid path transfer tube pipeline and culture solution go out Mouthful；Cell liquid flow path is successively including the hollow fibre filament outer space in cell liquid import, processing flow path, lumen import, lumen Between, lumen outlet, processing flow path, cell liquid outlet and optional cell fluid path transfer tube pipeline.

In certain embodiments, the processing flow path is machined or injection molding is arranged in the body interior.

In certain embodiments, the ontology of the media to cell culture modules is made of the upper, middle and lower, wherein：

It combines closely between upper layer and middle level, middle level and lower layer's binding face, no leakage；

The upper layer is equipped with culture supernatants import and culture supernatants and exports, respectively with the culture sap cavity that is set to middle level Connection；And connection culture supernatants import is exported with the processing flow path of middle level culture solution import with culture supernatants are connected to in The processing flow path of layer culture solution outlet；

The middle level is the frame structure of a middle part hollow out, which is equipped with and culture supernatants inlet communication Processing flow path, culture solution between middle level culture solution import, the import of culture sap cavity, middle level culture solution import and culture sap cavity import Chamber exports between middle level culture solution outlet, the outlet of culture sap cavity and the outlet of middle level culture solution of culture supernatants outlet Process processing flow path, the cell between flow path and cell liquid import, lumen import, cell liquid import and lumen import Processing flow path between chamber outlet, cell liquid outlet and lumen outlet and cell liquid outlet；

The hollow-out part of the culture sap cavity and lumen setting in middle level；With

The hollow fibre filament is placed in lumen, and both ends pass through the sealing block, is connected to respectively with culture sap cavity.

In certain embodiments, the media to cell culture modules further includes：

Culture fluid path transfer tube pipeline in the body exterior, the culture fluid path transfer tube pipeline connection culture are set Sap cavity is exported exports separated processing flow path with middle level culture solution；And/or

Cell fluid path transfer tube pipeline in the body exterior is set, described in the cell fluid path transfer tube pipeline connection Cell liquid import is exported with cell liquid.

In certain embodiments, the culture fluid path transfer tube pipeline and cell fluid path transfer tube pipeline are medical soft Pipe.

In certain embodiments, the upper layer is additionally provided with slot and chip pulling mouth, wherein the slot is slotting for positioning Enter culture solution module, the chip pulling mouth takes out culture solution module for facilitating.

In certain embodiments, sealing block is bonded with the contact surface leakproofness in middle level, and upper layer, lower layer are bonded with middle level Equal leakproofness fitting, the training is formed by the side wall of sealing block and upper layer inner wall, lower layer's inner wall and hollow-out part between face Nutrient solution chamber and lumen.

In certain embodiments, it is trained between the culture supernatants import and middle level culture solution import and on upper layer Nutrient solution is exported is arranged sedimeter between the outlet of middle level culture solution, with the pollution introduced to avoid puncture medical rubber stopper； Preferably, filter sizes are 0.2 μm.

The culture solution module of the utility model includes：

Closed ontology only by storing liquid import, the outlet of storage liquid is in communication with the outside；

It is located at intrinsic storage chamber, wherein the storage chamber and storage liquid import and storage liquid outlet；With

The ventilated membrane for allowing gas molecule to penetrate is used for sealed storage chamber.

In certain embodiments, the volume of the storage chamber is 0.6~2L, preferably 0.8~1.5L.

In certain embodiments, the ontology and ventilated membrane are transparent biocompatible materials.

In certain embodiments, storage chamber import, storage chamber outlet are all made of medical rubber stopper and are sealed.

In certain embodiments, the storage chamber with storage liquid inlet communication has storage chamber import, the storage chamber Import passes through default processing pipeline and the storage liquid inlet communication；The storage chamber with storage liquid outlet has storage Sap cavity exports, storage chamber outlet by default processing pipeline with the storage liquid outlet.

In certain embodiments, it is equipped with sedimeter after storage chamber outlet and storage chamber import, for filtering And obstruct impurity and bacterium enters in storage chamber.

In certain embodiments, the culture solution module is made of upper layer, middle level, lower layer and ventilated membrane, at the middle and upper levels Combine closely between middle level, middle level and lower layer's binding face, no leakage, at the middle and upper levels between middle level be equipped with ventilated membrane and/or Ventilated membrane is equipped between middle level and lower layer.

In certain embodiments, during storage liquid inlet and outlet and storage chamber inlet and outlet are all provided in middle level, when feed flow, adopts With hollow injection pin type bidirectional interface, the medical rubber stopper of direct penetration cell culture module and culture solution module simultaneously.

In certain embodiments, also there is filling exit, the interface to be sealed using medical rubber stopper in culture solution module, For the filling of initial incubation liquid；Preferably, sedimeter is further respectively had at filling exit.

The cell cultivation chip of the utility model contains media to cell culture modules as described herein and culture solution module, wherein institute The culture solution import and culture solution outlet for stating media to cell culture modules are exported and are stored with the storage liquid of the culture solution module respectively Liquid inlet communication.

The cell culture cabinet of the utility model, including：Casing is integrated in the fluid path driving in face in or on the cabinet Module, control module and the chip chamber for accommodating the cell cultivation chip described in claims 13, and optional be integrated in The thermal module and sensor assembly of cabinet inside；

Wherein, the fluid path drive module is used to that liquid to be driven to flow, including first driving means and the second driving device；

The control module includes：First control switch, for controlling first driving means；Second control switch, is used for Control the second driving device；Third control switch, for controlling gas conveying；It is switched with the 4th optional control, for controlling The heating or cooling of thermal module.

In certain embodiments, the chip chamber is independently of a container except cabinet, can be arranged on the container There are air admission hole and venthole, for being connected to venthole with the air admission hole on cabinet, to be connected to gas module；Or the core Piece chamber is a space in cabinet, the air admission hole and venthole that the space is only equipped with by cabinet after closing cabinet with it is outer Portion is connected to.

In certain embodiments, the thermal module includes semiconductor temperature-control piece and the temperature sense being mounted thereon member Part platinum resistance, for controlling heating or refrigeration；The sensor assembly includes temperature sensor, oxygen concentration sensor, titanium dioxide Concentration of carbon sensor and liquid pH value sensor.

In certain embodiments, it is additionally provided with front plate on the cabinet, display screen and control button is installed thereon, are used for The work operating condition of real-time display instrument and the control of instrument is operated.

In certain embodiments, it is also set on cabinet with air admission hole and venthole, for being connected to the gas module； With also integrated 1~5 gas control valve on case, it is turned on and off the third control switch control by respective numbers.

The cell culture apparatus of the utility model includes：Media to cell culture modules as described herein, culture solution as described herein The pipeline or circuit of module, fluid path drive module, gas module, thermal module, control module and each module of connection.

In one or more embodiments, the fluid path drive module includes：

First driving means, the circulation for controlling culture solution between culture solution module and media to cell culture modules；With

Second driving device, the circulation of the lumen inner cell liquid for controlling media to cell culture modules.

In certain embodiments, the communicating pipe in storage liquid outlet and culture solution import is arranged in the first driving means On the road, or it is arranged on connecting pipeline of the culture solution outlet with storage liquid import.

In certain embodiments, the media to cell culture modules includes external culture fluid path transfer tube pipeline, and described One driving device is arranged on the culture fluid path transfer tube pipeline.

In certain embodiments, second driving device is arranged on cell fluid path transfer tube pipeline.

In certain embodiments, the first driving means and the second driving device are clamp peristaltic pump, preferably Two-way peristaltic pump.

In certain embodiments, the gas module includes：Air inlet pipe, escape pipe, gas control valve and optional gas Body container and optional gas vortex mixer.

In certain embodiments, the gas container include accommodate compressed air, oxygen and carbon dioxide container, respectively Container is connected to gas vortex mixer respectively, and the air inlet pipe is connected to the gas vortex mixer.

In certain embodiments, it is equipped with filter device in the air inlet pipe and an air outlet pipe nozzle, gas is equipped in device Filter membrane.

In certain embodiments, connection accommodate compressed air, oxygen, carbon dioxide container and gas vortex mixer pipe Road is respectively equipped with gas control valve, and gas control valve is equipped with gas flowmeter.

In certain embodiments, the thermal module includes semiconductor temperature-control piece and the temperature sense being mounted thereon member Part platinum resistance, for controlling heating or refrigeration.

In certain embodiments, the control module includes：

First control switch, for controlling the first driving means；

Second control switch, for controlling second driving device；

Third control switch, for controlling gas conveying；With

4th control switch executes heating or refrigeration for controlling thermal module.

In certain embodiments, the control module further include for the entire cell cultivation process of real-time inspection and control, Realize the control circuit of automatic cytological culture.

In certain embodiments, the cell culture apparatus further includes sensor assembly, to real-time monitoring temperature, oxygen Gas concentration, gas concentration lwevel and pH value.

In certain embodiments, the sensor assembly includes that temperature sensor, oxygen concentration sensor, carbon dioxide are dense Spend sensor and liquid pH value sensor.

In certain embodiments, the media to cell culture modules, culture solution module and sensor assembly be positioned over equipped with into The closed chip intracavitary of stomata and venthole.

In certain embodiments, the integrated installation of the fluid path drive module, gas module, thermal module and control module In in cabinet, cabinet inside is equipped with the closed chip chamber for placing media to cell culture modules, culture solution module and sensor assembly, cabinet Outside is equipped with alternating interface between man and computer, control button.

In certain embodiments, the culture solution is the special serum free medium of immunocyte；

In certain embodiments, the cell is selected from peripheral blood mononuclear cells, neural stem cell or tumour cell.

In certain embodiments, the material contacted with cell in described device uses disposable biological compatibility material Material, and by disinfection and sterilization treatment.

The utility model also provides a kind of box set, which includes media to cell culture modules and culture solution mould as described herein Block, or contain chip as described herein.

In certain embodiments, the box set, which further includes integrated installation, fluid path drive module described herein, gas control The cabinet and batch cultur consumptive material of valve processed, thermal module and control module, the medical disposable material include：It is as described herein Media to cell culture modules, culture solution module and all pipelines.

Description of the drawings

Fig. 1 is the planar structure schematic diagram of chip type immunocyte culture systems provided by the utility model.

Fig. 2 is the structural schematic diagram of immunocyte culture chip provided by the utility model.

Fig. 3 is the structural schematic diagram of the cell culture unit of the utility model culture chip.

Fig. 4 is the structural blast schematic diagram of the cell culture unit of the utility model culture chip.

Fig. 5 is the structural schematic diagram of one doughnut chunking example of the utility model.

Fig. 6 is the structural blast schematic diagram of the culture solution unit of the utility model culture chip.

Fig. 7 is the overall structure diagram of the utility model cell culture apparatus.

Fig. 8 is the overall structure diagram that culture chip is put into the utility model cell culture apparatus.

Specific implementation mode

The utility model is illustrated below in conjunction with specific implementation mode.It should be understood that these embodiments are merely to illustrate this Utility model rather than limitation the scope of the utility model.Other aspects of the utility model due to this disclosure, It is obvious to those skilled in the art.

The cell culture system of the utility model is chip type immunocyte culture systems (device).The utility model it is thin Born of the same parents' culture apparatus can realize a kind of new immunocyte culture scheme, can every clinical patients sample of independent process cell training It supports.By taking out a certain amount of cell, especially immunocyte from patient's body, it is put into the culture systems of the utility model and trains Support, the aim cell of final available respective numbers, as DC cells Antigen culture cell scale 0.5~2 × 10⁸ Sum on, the cell scale of the quantity amplification cultivation of T cell is 0.5~2 × 10⁹Sum.Entire immunocyte culture Journey can automation control, it is only necessary to regularly replace culture solution module, cell culture carries out (no opening in the environment of closing always Formula operates, and effectively third party is avoided to pollute), the immunocyte finally recycled may be directly applied to clinic, feed back patient's body Carry out immune cell therapy.Entire incubation builds the training method of class blood vessel by hollow fibre filament, and filament is built in In lumen, culture solution flows into other side culture chamber by filament from side culture sap cavity, and it is certain that cell is given in flowing Shearing force, and ensure that cell intracavity liquid also has certain mobility, 3D to cultivate the life for effectively simulating immune cell growth Substance environment also improves the efficiency of culture solution exchange.

This cell culture apparatus includes media to cell culture modules, culture solution module, fluid path drive module, gas module, temperature It is connected between module and six big main modular of control module and culture solution module, media to cell culture modules, fluid path drive module Pipeline.In certain embodiments, this cell culture apparatus further includes sensor assembly.Fig. 1 shows Tthe utility model system A structural schematic diagram, including chip chamber 1, media to cell culture modules 2, culture solution module 3, fluid path drive module 4, gas module 5, thermal module 6, sensor assembly 7 and control module 8.

Media to cell culture modules

Media to cell culture modules is the place of cell culture, including culture sap cavity and two big structure of lumen, is led between the two Sealing block is crossed to be separated by.Culture sap cavity is to accommodate the place of fresh medium or the culture solution through exchange, and lumen is cell Grow place.Hollow fibre filament is provided in lumen, filament inner spout both ends are connected to culture sap cavity.It cultivates in sap cavity Fresh medium passes through lumen in hollow fiber conduit pipe.By the micropore on hollow fibre filament, lumen and culture sap cavity Between can carry out small molecule exchange, take away cell growth generation metabolic waste object, supplement cell needed for nutriment.

More specifically, media to cell culture modules includes：Only by culture solution import, culture solution outlet, cell liquid import and Cell liquid exports the closed ontology being in communication with the outside；It is located within the body the culture sap cavity and lumen in portion, wherein culture sap cavity Including it is independent respectively with the culture sap cavity of culture solution inlet communication and with the culture sap cavity of culture solution outlet, lumen with Cell liquid import and cell liquid outlet；Sealing block, for being separated to form lumen and culture sap cavity；Be placed in lumen It is interior, have on wall do not allow cell by micropore hollow fibre filament；Wherein, hollow fibre filament passes through sealing block, hollow Filametntary both ends are connected to the culture sap cavity of culture solution input end, with the culture sap cavity of culture solution outlet end respectively.Cell All building blocks of module are cultivated, including the material of ontology and sealing block are all biocompatible materials, it is preferably transparent Material.

Culture solution import and culture solution outlet are two openings being located on media to cell culture modules, are used for and culture solution module Connection, the inlet and outlet flowed in and out for culture solution.Lumen both sides can be located at by cultivating sap cavity, be separated by sealing block, Hollow fibre filament passes through lumen to be connected to the culture sap cavity of lumen both sides；Alternatively, culture sap cavity is located at lumen the same side, It is separated by sealing block between two culture sap cavities and between culture sap cavity and lumen.With the training of culture solution inlet communication Nutrient solution chamber itself is equipped with culture sap cavity import, the culture sap cavity import and the culture solution inlet communication.It exports and connects with culture solution Logical culture sap cavity itself is equipped with the outlet of culture sap cavity, culture sap cavity outlet and culture solution outlet.Cultivate sap cavity import It is each provided in ontology with the outlet of culture sap cavity, is connected respectively with culture solution import and culture solution outlet by intrinsic processing flow path It is logical.Culture solution import and culture solution outlet can be used medical rubber stopper and be sealed, rear side (i.e. inside, close to the one of culture sap cavity Side) it can be respectively equipped with sedimeter, it, can when culture solution inlet and outlet being avoided to be connected to culture solution module to broth filtrate The pollution that can be introduced.In general, the total measurement (volume) of culture sap cavity is 0.2~0.5L, preferably 0.3~0.35L.

In certain embodiments, media to cell culture modules further includes the culture fluid path transfer tube pipeline for being located at body exterior, It cultivates and is disconnected among the processing flow path between sap cavity outlet and culture solution outlet, by the culture fluid path transfer tube pipeline connection. In this case, the processing flow path of the disconnection forms two interfaces on the body, connects to be connected to culture fluid path transfer tube pipeline It is logical.

Cell liquid import and cell liquid outlet be located on media to cell culture modules another two opening, for cell liquid flow into and The inlet and outlet of outflow.There is lumen itself lumen import and lumen to export, respectively with cell liquid import and cell Liquid outlet.The outlet of lumen import and lumen is each provided at body interior, can be by being located at the processing flow path of body interior Respectively with cell liquid import and cell liquid outlet.Media to cell culture modules further includes the cell fluid path driving for being located at body exterior Pump conduit, the cell fluid path transfer tube pipeline are used for being connected to cell liquid import and are exported with cell liquid.The volume of lumen be 0.4~ 1L, preferably 0.6~0.8L.

Hollow fibre filament is intermediate throughholes filament, and the length of 130~170mm, 20~50 μm of thickness of pipe wall is covered with hole on wall Diameter is less than 1 μm of micropore, to prevent cell from passing through.Small-molecule substance or dissolved oxygen can pass through the micropore and fiber on filament wall Lumen outside silk carries out substance and gas exchanges, and the validity of culture solution in lumen, realization is kept to continue cell Nutrient.The material of hollow fibre filament can be biocompatible materials commonly used in the art, preferably biocompatibility compared with Good polypropylene, polysulfones, polypropylene be fine or polyethylene etc..The quantity of hollow fibre filament in lumen is had no it is specifically limited, but Under normal conditions, it should make the space (volume) in lumen outside hollow fibre filament in the range of 0.2~0.6L.

In general, the media to cell culture modules of the utility model has culture solution flow path and cell liquid flow path.Culture solution flow path is The access of culture solution flowing, include successively culture solution import, processing flow path, culture sap cavity import, input end culture sap cavity, in Inside hollow fiber silk, the culture sap cavity of outlet end, the outlet of culture sap cavity, process flow path, optional culture fluid path transfer tube pipeline It is exported with culture solution.Cell liquid flow path is the access of cell liquid flowing, includes cell liquid import, processing flow path, lumen successively The hollow fibre filament external space, lumen outlet, processing flow path, cell liquid outlet in import, lumen and the driving of cell fluid path Pump conduit.Culture solution flow path can form circulation loop with culture solution module as described herein, and in cell liquid flow path, cell fluid path It drives pump conduit with after cell liquid inlet communication, forms circulation loop.

For example, when Cyclic culture liquid, culture solution is flowed out from culture solution module, enters cell culture mould from culture solution import Block flows to culture sap cavity import via processing flow path, flows into the culture sap cavity of culture solution input end, flow through in hollow fibre filament Empty inner cavity reaches the culture sap cavity of culture solution outlet end, goes out via culture sap cavity outlet stream, into processing flow path, is then out Culture solution exports, and flows back into culture solution module, a circulation loop, herein also referred to as culture solution circulation loop is consequently formed.At certain In a little embodiments, disconnected among the processing flow path between the outlet of culture sap cavity and culture solution outlet, by being placed outside cell culture Culture fluid path transfer tube pipeline connection except module body.And cell liquid then the hollow fibre filament external space in lumen, It is recycled between cell liquid import and cell liquid outlet, forms cell liquid circulation loop.

Herein, processing flow path can be arranged through the techniques such as being molded or machining in the body interior；Cultivate fluid path driving Pump conduit and cell fluid path transfer tube pipeline can be conventional flexible pipe for medical purpose.

In certain embodiments, media to cell culture modules is by the upper, middle and lower up of three layers.Upper layer and middle level, in Leakproofness is bonded between layer and lower layer binding face, and no leakage is air tight.

Upper layer is equipped with culture supernatants import and culture supernatants export, and media to cell culture modules is flowed into and out for culture solution Culture solution inlet and outlet；And connection culture supernatants import and the processing flow path of middle level culture solution import be connected to upper layer Culture solution exports and the processing flow path of middle level culture solution outlet.Upper layer can also be equipped with slot and chip pulling mouth.Slot can be used for fixed Culture solution module is inserted into position, and chip pulling mouth, which can be used for facilitating, takes out culture solution module.In certain embodiments, culture supernatants Import, culture supernatants outlet can be used medical rubber stopper and be sealed.

It is 5~15mm, preferably 8~10mm that middle level, which has certain thickness, usual thickness, is the frame knot of a middle part hollow out Structure, the frame structure are equipped with middle level culture solution import, the import of culture sap cavity, middle level culture with culture supernatants inlet communication It trains in the middle level of processing flow path, the outlet of culture sap cavity and culture supernatants outlet between liquid import and culture sap cavity import Nutrient solution outlet, culture sap cavity outlet middle level culture solution export between processing flow path and cell liquid import, lumen import, Processing flow path, lumen outlet, cell liquid outlet and lumen outlet between cell liquid import and lumen import and cell Processing flow path between liquid outlet.Between culture supernatants import and middle level culture solution import and culture supernatants outlet and Middle level can be respectively equipped with filter device between culture solution outlet, to broth filtrate, avoid culture sap cavity inlet and outlet and culture The pollution that liquid module may introduce when being connected to.Filter sizes are preferably 0.2 μm of aperture or following.

The part of hollow out is for accommodating doughnut chunking in middle level.Herein, doughnut chunking is by hollow fibre filament Be arranged hollow fibre filament both ends sealing block composition.Hollow fibre filament pass through the sealing block, respectively with culture solution The culture sap cavity of input end and the culture sap cavity connection of culture solution outlet end.In general, the both ends of hollow fibre filament do not surpass respectively Go out the outside of sealing block, is preferably flushed with the outside of sealing block (side i.e. towards culture sap cavity).Sealing block with The hollow fibre filament outer wall leakproofness fitting passed through.Meanwhile sealing block is bonded with the contact surface in middle level also leakproofness.When After leakproofness is bonded between upper layer, lower layer and middle level binding face, by sealing block and upper layer inner wall, lower layer's inner wall and hollow-out parts The side wall divided forms the culture sap cavity and lumen.

Middle level culture solution import can by be built in upper layer processing flow path and upper layer culture solution inlet communication, middle level training Nutrient solution outlet can by be built in upper layer processing flow path and upper layer culture solution outlet.Middle level culture solution import and middle level Culture solution outlet can be connected to the culture sap cavity of inlet end and an outlet end respectively by the processing flow path set on body interior.At certain In a little embodiments, disconnected among the processing flow path between the outlet of culture sap cavity and the outlet of middle level culture solution, it is upper in middle level to be formed Two interfaces, the culture fluid path transfer tube pipeline connection external by being placed in media to cell culture modules sheet.

Cell liquid import is via the processing flow path that is located inside middle level and lumen inlet communication, and cell liquid outlet is via setting Internal processing flow path and lumen outlet in middle level, are finally connected to the hollow fibre filament external space of lumen, and thin Pass through cell fluid path transfer tube pipeline connection between cytosol import and cell liquid outlet.

When carrying out culture solution cycle, culture solution is entered by culture supernatants import, passes through the processing being built in upper layer Flow path flows into middle level culture solution import through filter device, then processes the culture solution that flow path enters culture sap cavity input end by middle level Chamber flows through the hollow cavity of hollow fibre filament, and into the culture sap cavity of outlet end, processed flow path is exported by culture sap cavity, according to The culture solution outlet of secondary outflow middle level and culture supernatants outlet, flow back to culture solution module, and closed culture solution cycle is consequently formed Circuit.When media to cell culture modules is equipped with culture fluid path transfer tube pipeline, the outlet of culture solution outflow culture sap cavity stayed one section to add After work flow path, into the culture fluid path transfer tube pipeline, processing flow path is passed through, sequentially enters middle level culture solution outlet and upper layer Culture solution exports, and flows back to culture solution module, closed culture solution circulation loop is consequently formed.Doughnut group is flowed through in culture solution When block, nutritional ingredient is flowed to by the micropore on hollow fibre filament wall outside hollow fibre filament, is located at outside doughnut to nourish Cell.And cell liquid then the processing flow path between cell liquid import, cell liquid import and lumen import, lumen import, The hollow fibre filament external space of lumen, lumen outlet, lumen export processing flow path, cell liquid between cell liquid outlet Cycle in outlet and cell fluid path transfer tube pipeline.

Fig. 2, Fig. 3, Fig. 4 and Fig. 5 respectively show one media to cell culture modules example (media to cell culture modules of the utility model 2) overall structure and Each part.Media to cell culture modules 2 includes upper layer 201, middle level 203 and lower layer 204, all with biology Compatibility material manufactures.It combines closely between three layers of binding face, binding site uses effective binder or ultrasonic bonding It is combined together, with liquid-leakage preventing and/or gas leakage.

Upper layer 201 includes culture supernatants import 205, culture supernatants outlet 206, slot 207 and chip pulling mouth 208. Culture supernatants import 205 and culture supernatants outlet 206 pass in and out the inlet and outlet of media to cell culture modules 2 for culture solution.Slot 207 It is inserted into culture solution module for positioning.Culture solution import 205 and culture solution outlet 206 in can be respectively equipped with filter device 205b and 206b.The filter device is sedimeter, and built-in filter membrane can be used for avoiding puncturing the pollution that medical rubber stopper may introduce. Chip pulling mouth 208, which can be used for facilitating, takes out culture solution module.

Middle level 203 is the frame structure of the middle part hollow out with certain thickness, which cultivates equipped with middle level Liquid import 211, culture sap cavity import 211a, middle level culture solution outlet 212, culture sap cavity outlet 212a, the outlet of culture sap cavity The processing stream between processing flow path 9a, cell liquid import 215 and lumen import 215a between 212a and middle level culture solution outlet 212 Road 10a, culture fluid path transfer tube pipeline 213, cell fluid path transfer tube pipeline 214, lumen import 215a, cell liquid import 215, lumen outlet 216a and cell liquid outlet 216.In middle level 203 shown in Fig. 4, culture sap cavity outlet 212a is trained with middle level Processing flow path between nutrient solution outlet 212 is disconnected, and two openings are formed on outer wall in middle level, is driven for connecting culture fluid path The both ends of dynamic pump conduit 213.

Media to cell culture modules 2 further includes the doughnut chunking 202 of 203 hollow-out part of embedded middle level.Fig. 3 and 4 is shown Illustrative 202 structure of doughnut chunking.The doughnut chunking 202 includes more hollow fibre filaments 209 placed side by side With the sealing block 210 positioned at 209 both ends of hollow fibre filament.Sealing block 210 can be by any appropriate biocompatible materials It is made.Every 209 both ends of hollow fibre filament are each passed through sealing block 210, and part is passed through to be bonded with sealing block leakproofness.It is logical Often, hollow fibre filament flushes on the outside of the part of sealing block and sealing block.Every piece of sealing block connects with middle level 203 Contacting surface leakproofness is bonded.There are certain spaces between middle level 203 respectively at the both ends of sealing block 210, to form culture sap cavity 12 and culture sap cavity 14.It is the lumen 13 for accommodating the hollow fibre filament 209 between culture sap cavity 12 and 14.

Fig. 5 shows the structure of another doughnut chunking 202a, including hollow fibre filament 209 and sealing block 210. Hollow fibre filament 209 is in 180 ° of U-bends, therefore two culture sap cavities are located at the same side of lumen.Similarly, two cultures Sap cavity still has sealing block to separate, which can be integrally formed with sealing block 210, or independent sealing gear Block, but need two culture sap cavities can be separated, ne-leakage.It, can be as mentioned before when using doughnut chunking shown in fig. 5 The other structures of media to cell culture modules, including each inlet and outlet and each processing flow path are set.

In general, by taking Fig. 4 as an example, culture solution is entered by culture supernatants import 205, by being built in adding in upper layer 201 The middle level culture solution import 211 in work fluid communication middle level 203 enters culture by processing flow path via culture sap cavity import 211a Sap cavity 12 flows through the hollow cavity of hollow fibre filament 209, and into culture sap cavity 14, culture is flowed through from culture sap cavity outlet 212a Fluid path drives pump conduit 213, into processing flow path 9a, finally by middle level culture solution outlet 212 and culture supernatants outlet 206 Culture solution module 3 is flowed back to, a closed culture solution circulation loop 9 is consequently formed.Hollow fibre filament 209 is flowed through in culture solution When, nutritional ingredient is flowed to by the micropore on hollow fibre filament wall outside hollow fibre filament, is located at outside hollow fibre filament to nourish Cell.

And cell liquid circulation loop 10 is then formed outside hollow fibre filament, this circulation loop is by 13 doughnut of lumen Space, the processing flow path 10a that is connected to the space outside silk and the composition of cell fluid path transfer tube pipeline 214.Cell liquid is by thin Cytosol import 215 enters, and flows through after processing flow path 10a through entering 13 hollow fibre filament outer space of lumen by lumen import 215a Between, then by lumen outlet 216a outflows, via processing flow path outflow cell liquid outlet 216, into cell fluid path transfer tube Pipeline 214, then cell liquid import 215 is flowed into, form the circulation loop 10.

Entire incubation builds the training method of class blood vessel by hollow fibre filament, and filament 209 is built in lumen In 13, culture solution flows into other side culture chamber 14 by filament 209 from side culture sap cavity 12, and it is certain that cell is given in flowing Shearing force, and ensure cell intracavity liquid also have certain mobility, effectively simulate the biocycle of immune cell growth Border also improves the efficiency of culture solution exchange.

Culture solution module

Culture solution module includes mainly storage cavity configuration, accommodates fresh cell culture fluid for storing, can be primary The biological reaction container of property, such as disposable biological reaction bag or biometric storage chip.Culture solution module is mainly used for training to cell It supports module and culture solution is provided, tool is the outlet of storage liquid there are two interface, wherein first interface, for the training with media to cell culture modules Nutrient solution inlet communication；Second interface is storage liquid import, for the culture solution outlet with media to cell culture modules.Storage chamber Volume is 0.6~2L, preferably 0.8~1.5L.

In certain embodiments, there is ventilated membrane, the ventilated membrane gas-permeable, liquid-impermeable to allow film outer gas in culture solution module Body molecule is dissolved in by the film in the culture solution in storage chamber, to ensure the dissolved oxygen amount of culture solution.

In certain embodiments, the culture solution module is made of the upper, middle and lower three, and upper layer is in Leakproofness is bonded between layer, middle level and lower layer binding face, no leakage.Gripped between upper layer and middle level ventilated membrane and/or middle level with Ventilated membrane is gripped between lower layer.Preferably, upper, middle and lower-ranking and ventilated membrane all use transparent biocompatible materials system It makes.Storage liquid inlet and outlet are arranged in middle level, are connected to respectively with the storage chamber in middle level inlet and outlet by default processing pipeline.It deposits Liquid storage import, the outlet of storage liquid are all made of medical rubber stopper and are sealed.Hollow injection pin type bidirectional interface is used when feed flow, directly The medical rubber stopper of penetration cell culture module and culture solution module simultaneously.In certain embodiments, storage chamber outlet and storage Chamber entrance further respectively has sedimeter.

In certain embodiments, also there is filling exit, it is close which can be used medical rubber stopper progress in culture solution module Envelope, can be used for the filling of initial incubation liquid.Preferably, sedimeter is further respectively had at filling exit.

Gas can be entered by the ventilated membrane in culture solution module in cell culture fluid, and ventilated membrane can provide the gas of large area Liquid contacts, and higher oxygen mass transfer coefficient can be provided under lower gas flow rate.

Fig. 2 and Fig. 6 show respectively one culture solution module example (culture solution module 3) of the utility model overall structure and Each part.Culture solution module 3 includes upper layer 301, middle level 303 and lower layer 304, is all manufactured with biocompatible materials. Ventilated membrane 302 is designed between upper layer 301 and middle level 303, middle level 303 and lower layer 304, between three layers and layer block and ventilated membrane Between combine closely to form storage chamber 305, binding site is combined together using effective binder or ultrasonic bonding, nothing The case where leakage and/or gas leakage.Culture solution module 3 has storage liquid outlet 306 and storage liquid import 307, passes through default processing Pipeline connection is set to the storage chamber outlet 306a and storage chamber import 307a in middle level 303.When feed flow, using hollow injection pin type Bidirectional interface 311 directly penetrates connection storage liquid outlet 306 and culture solution import 205,307 and of storage liquid import simultaneously respectively The sealing medical rubber stopper of culture solution outlet 206, carries out fluid infusion, while also ensuring good closure.Storage chamber outlet 306a, Sedimeter 309 is equipped at storage chamber import 307a.3 middle level 303 of culture solution module further includes filling exit 310, for just Beginning culture solution it is filling, sedimeter 309 also is provided at filling exit 310.

Culture solution module can be positioned over the top of media to cell culture modules or is inserted by the slot 207 of media to cell culture modules solid This fixed module, chip pulling mouth 308 are convenient for the installation or taking-up of module.

Fluid path drive module

Fluid path drive module is for driving liquid to be continued to flow into stable flow velocity and flowing out media to cell culture modules and/or training Nutrient solution module, to ensure the circulation of flow path in whole system.Fluid path drive module usually contains the device of driving liquid flowing, Preferably clamp peristaltic pump.It is further preferred that peristaltic pump is two-way peristaltic pump.In general, fluid path drive module includes two drivings Device is located on culture solution circulation loop and cell liquid circulation loop, the flowing and cell liquid for driving culture solution Flowing.

As shown in Figure 1, fluid path drive module 4 includes first driving means 401 and the second driving device 402.First driving Device 401 is located on culture solution circulation loop 9, and the second driving device 402 is located on cell liquid circulation loop 10.

As shown in figure 8,401 clamping of first driving means peristaltic pump is fixed on culture fluid path transfer tube pipeline 213, for controlling The circulation of culture solution between media to cell culture modules and culture solution module processed；Second driving device peristaltic pump, 402 clamping is fixed on carefully Cytosol road drives pump conduit 214, for realizing the cycle of cell liquid.

In general, the rotating speed of peristaltic pump is normally controlled in the range of 10~50r/min, such as 20~30r/min.

Gas module

Gas module is mainly used for conveying mixed gas into chip chamber 1, the mixed gas usually contain air, oxygen and Carbon dioxide etc..Wherein oxygen is necessary to cell metabolism, and ventilatory capacity is needed with bioreactor (i.e. cell culture mould Block) volume and cell concentration raising and increase, but higher oxygen content can also generate cell toxicity, therefore oxygen concentration meeting Accurate control is carried out by sensor.

In general, gas module includes air inlet pipe and an air outlet pipe, air inlet pipe one end is used to be connected to the air admission hole of culture apparatus, The other end is then used to be connected to gas container；Escape pipe with the venthole of culture apparatus for being connected to.Air inlet pipe is equipped with gas Control valve is turned on and off control conveying of the gas to culture apparatus by gas control valve.In air inlet pipe and an air outlet pipe also It can be equipped with filter device, it is interior to be equipped with gas filter membrane.Filter device in air inlet pipe is generally arranged at gas control valve and is filled with culture It sets between air inlet nose end.

When needing to convey gas, air inlet pipe one end is connect with gas container, gas control valve is opened, needed for conveying Gas.

Gas can be the gas being pre-mixed.In this case, air inlet pipe one end can directly be mixed with containing this in advance The container of the gas got togather connects.

In certain embodiments, gas is not pre-mixed.In this case, gas module further includes vortex mixer.Mixing Device is located between the gas control valve of air inlet pipe and filter device (if any), and is respectively provided with gas to be mixed (such as compressed air, oxygen Gas and carbon dioxide) gas container be connected.Gas control valve is set respectively on the access of each gas container and vortex mixer, passes through gas Body control valve is turned on and off conveying of the gas to vortex mixer in each gas container of control.Therefore, the quantity of gas control valve It can be 1~5, such as 1~3.

In certain embodiments, gas module further includes 1 or several gas containers, and such as 1~5 or 1~5 gas holds Device.

Fig. 1 and Fig. 8 shows one gas module example of the utility model.Specifically, gas module 5 includes air inlet pipe 501 and escape pipe 507, wherein 501 one end of air inlet pipe be connected to the air admission hole into chip chamber 1, the other end is connected to vortex mixer 502, Compressed air 504, oxygen 505 are connected to carbon dioxide 506 in gas vortex mixer 502.Air inlet pipe 501 and escape pipe 507 Nozzle is respectively arranged with filter device 508, and gas filter membrane is housed in device.Control module 8 controls third control switch and compresses respectively Air, oxygen, carbon dioxide gas control valve 503 open and close, make each gas in gas vortex mixer 502 after mixing Into chip chamber.When there are multiple gas control valves 503, the third control switch of respective numbers can be set.

Thermal module

Thermal module is for ensureing that entire chip intracavitary maintains suitable cell culture temperature.The main body of thermal module is Semiconductor temperature-control piece can be heated or be freezed, and persistently be confessed with stable heat dissipation capacity.Preferably, semiconductor temperature-control on piece Temperature sensitive member platinum resistance is installed.As illustrated in figures 1 and 8, thermal module 6 is installed on the inside of chip chamber 1, by control module 8 the 4th control switch controls.

Control module

Control module includes the first control switch, the second control switchs, third control switch and the 4th controls switch.First Control switch for controlling the driving device between culture solution module and media to cell culture modules, via first control switch open with Close the circuit for making the media to cell culture modules, drive module and culture solution module be formed with or without cycle, or control culture The flow velocity or flow of liquid；Second control switch is for controlling the between the import of media to cell culture modules cell liquid and cell liquid outlet Two driving devices make to be formed with or without between the cell culture import and outlet via the Push And Release of the second control switch The circuit of cycle, or control the flow velocity or flow of cell liquid；Third control switch, for controlling gas module to device chip chamber The conveying of gas；4th control switch, the temperature liter or temperature drop of the semiconductor chip for controlling thermal module.

Control module may also include corresponding control circuit, and for controlling the first above-mentioned control switch, the second control is opened It closes, third control switch and the 4th control switch.In general, the first control switch is for controlling first driving means 401, the second control System switch is for controlling the second driving device 402；Third control switch, for controlling gas control valve 503；4th control is opened It closes, the semiconductor chip 6 for controlling thermal module.Control module further includes display, and driver is for showing described device According to the environment numerical value in scheduled program operating condition and real-time acquisition device, the operating status of device is shown.Display can To be touch screen displays.Alternatively, control module may include control panel, including control button, to control the first control Switch, the second control switch, third control switch and the 4th control are switched on and are closed.Or system control module may include Both display and control button.By control module, utility model device accurately controls entire cell cultivation process, real Existing full-automation immunocyte culture.

Sensor assembly

In certain embodiments, the cell culture system of this paper further includes sensor assembly.More specifically, the sensing Device module is used to detect the environmental parameter in cell culture system, including but not limited to temperature, oxygen concentration, gas concentration lwevel With liquid pH value etc..

The exemplary sensor assemblies 7 for having put on display this paper cell culture systems of Fig. 1 and Fig. 8, including temperature sensor 701, Oxygen concentration sensor 702, gas concentration lwevel sensor 703 and liquid pH value sensor 704.Temperature sensor 701 is used for Temperature at detection culture chip position；Oxygen concentration sensor 702 is used to monitor the concentration of oxygen gas in chip chamber 1；Two Carbonoxide concentration sensor 703 is used to monitor the concentration of carbon dioxide gas in chip chamber 1；Liquid pH value sensor 704 is non- Touch sensor, for monitoring liquid pH value in culture chip.

Cultivate chip

The utility model also provides a kind of culture chip, and the culture chip is by media to cell culture modules as described herein and culture Liquid module forms, and the two passes through respective orifice.Specifically, the culture solution import of media to cell culture modules and culture solution go out Mouth exports with the storage liquid of culture solution module respectively and stores liquid inlet communication, such as two-way by hose or hollow injection pin type Orifice.

For example, Fig. 2 shows the structural schematic diagram of the utility model culture chip, including media to cell culture modules 2 and culture Liquid module 3, culture solution module 3 are inserted into media to cell culture modules 2 via the slot on media to cell culture modules 2.Culture solution module is deposited Liquid storage exports the culture solution inlet communication with media to cell culture modules, and the culture solution outlet for storing liquid import and media to cell culture modules connects It is logical, form a complete culture chip.

Cell culture cabinet

A kind of cell culture cabinet is also provided herein, including：Casing is integrated in this paper institutes in face in or on the cabinet Fluid path drive module, control module and the chip chamber stated.In certain embodiments, which further includes being integrated in The thermal module and sensor assembly of cabinet inside.

Cabinet can be the size and shape of any suitable size, can be made of any suitable material.

Fluid path drive module includes first driving means and the second driving device.Driving device is arranged in liquid as described herein Can be clamp peristaltic pump, preferably bidirectional peristaltic pump on body circulation circuit.

Control module may include：First control switch, for controlling first driving means；Second control switch, for controlling Make the second driving device；Third control switch, for controlling gas conveying；It is switched with the 4th control, for controlling thermal module Heating or cooling.Third control switch can be set according to the quantity of gas control valve, for example, 1~5 or 1~3 third control System switch.

Chip chamber can be independently of a container except cabinet, by stainless steel and other metal materials or transparent organic glass Equal materials are made；Can also be a space in cabinet, the air inlet that the space is only equipped with by cabinet after closing cabinet Hole and venthole are connected to outside.Chip intracavitary can be equipped with suitable fixed structure, convenient for being fixed at the module of intracavitary, such as Media to cell culture modules, culture solution module, thermal module and sensor assembly etc..It, can be in core when chip chamber is independent container Suitable fixed structure is set outside piece chamber and/or in cabinet, which is fixed in cabinet.When chip chamber is independent Container when, may be provided with air admission hole and venthole on the container, for being connected to venthole with the air admission hole on cabinet, with Gas module is connected to.When the space that one that chip chamber is cabinet can be sealed, thermal module and sensor assembly usually collect At in cabinet.

Thermal module includes semiconductor temperature-control piece and the temperature sensitive member platinum resistance that is mounted thereon, for controlling heating Or refrigeration.Sensor assembly includes temperature sensor, oxygen concentration sensor, gas concentration lwevel sensor and liquid pH value sensing Device.

It can also be equipped with front plate on cabinet, display screen and control button can be installed thereon, be used for the work of real-time display instrument Make operating condition and the control of instrument is operated.

It can be also set on cabinet with air admission hole and venthole, for being connected to gas module as described herein.In certain implementations Can be also integrated in scheme, on cabinet with one or several (such as 1~5 or 1~3) gas control valves, be turned on and off by The third control switch control of respective numbers.

Fig. 7 shows the schematic diagram of the utility model cell raise machine case portion separation structure.The cell culture cabinet 101 of Fig. 7 Including chip chamber 1, window 102, incubator overhead door 103, heat emission hole 105, first driving means 401, the second driving device 402, display screen 801 and control button 802.

Cell culture apparatus

The cell culture apparatus of the utility model includes that media to cell culture modules as described herein, culture solution module, fluid path are driven Dynamic model block, gas module, thermal module and control module and culture solution module, media to cell culture modules, fluid path drive module it Between the pipeline that connects.In certain embodiments, cell culture apparatus further includes sensor assembly.In the device, cell culture Culture solution import and the culture solution outlet point that module and culture solution module pass through respective orifice, such as media to cell culture modules It is not exported with the storage liquid of culture solution module and stores liquid inlet communication, such as pass through hose or hollow injection pin type bidirectional interface Connection.

In general, fluid path drive module as described herein, the gas control valve in gas module, thermal module, sensor die Block and control module can integrated installation in the cabinet of cell culture apparatus, display, control button are installed outside cabinet.Carefully Born of the same parents, which cultivate module and culture solution module, can then be placed in the chip intracavitary of cabinet.

In certain embodiments, media to cell culture modules, culture solution module, thermal module and sensor assembly are placed in core Piece intracavitary.In these embodiments, fluid path drive module, the gas control valve of gas module and control module are integrated in cabinet It is interior.

Chip chamber is the subenvironment space for carrying out cell culture, the enclosed environment for building culture chip, convenient for control Cultivate the temperature and gas member condition of chip environments.Media to cell culture modules, culture solution module and its phase can be put into chip chamber The connecting pipeline of pass.Thermal module and sensor assembly are also mountable to chip intracavitary.Air admission hole can be set and gone out on chip chamber Stomata, for being connected to gas module.

Chip chamber can be an independent container, by materials systems such as stainless steel and other metal materials or transparent organic glass At；Can also be cell culture apparatus cabinet in a space, after closing cabinet only by air admission hole and venthole with Outside connection.Chip intracavitary can be equipped with suitable fixed structure, convenient for being fixed at the module of intracavitary, such as cell culture mould Block, culture solution module, thermal module and sensor assembly etc..It, also can be outside chip chamber when chip chamber is independent container And/or the suitable fixed structure of setting in cabinet, to be fixed in cabinet.When chip chamber is independent container, the appearance Air admission hole and venthole are may be provided on device, for being connected to venthole with gas module by the air admission hole on cabinet.

Fig. 1 shows the structural schematic diagram of the utility model cell culture apparatus, including chip chamber 1, media to cell culture modules 2, culture solution module 3, fluid path drive module 4, gas module 5, thermal module 6, sensor assembly 7 and control module 8.

Fig. 8 illustrates the overall structure and internal structure of cell culture apparatus.It is inserted into the cell culture apparatus 101 of Fig. 8 Culture chip, including chip chamber 1, media to cell culture modules 2, culture solution module 3, gas module 5, thermal module 6, control module 8, sensor module 701-704, first driving means 401, the second driving device 402, display screen 801 and control button 802 with And overhead door operator 104.Gas module 5 includes air inlet pipe 501, escape pipe 507, compressed air container 504, oxygen container 505, two Aoxidize carbon vessel 506, gas vortex mixer 502, gas control valve 503 and filter device 508.Fig. 8 also shows circulation loop 9 With 10.Incubator overhead door 103 is put into or takes out culture chip when opening, form accommodating for sealing when closing and cultivate chip Chip cavity space.Overhead door operator 104 is for controlling opening or closing for incubator overhead door 103.Heat emission hole 105 is used for cell culture The heat dissipation of case internal component.The display screen 801 and control button 802 installed in cabinet front plate, can real-time display instrument The operating condition that works and the control of instrument is operated.

Box set

The utility model also provides a kind of box set, which includes media to cell culture modules and culture solution mould as described herein Block, or including culture chip as described herein.It may also include connecting tube in box set, such as formation culture fluid path transfer tube pipeline and carefully Cytosol road drives the connecting tube of pump conduit.The media to cell culture modules, culture solution module or the culture chip and connecting tube can be with All it is medical disposable material.It is dismountable between connecting tube and media to cell culture modules, culture solution module.

It may also include cell culture cabinet as described herein in box set, which can be integrated with except cell as described herein is trained Support all other component outside module and culture solution module, the including but not limited to gas in fluid path drive module, gas module Control valve, thermal module, sensor assembly and control module.If it is desired, be also provided in box set connecting tube (snorkel) and Optional gas vortex mixer can be equipped with filter device on tracheae, inside set gas filter membrane.

But in certain embodiments, when chip chamber is independently of a container except cabinet, it is preferable that in cabinet The gas control valve and control module being only integrated in fluid path drive module, gas module, wherein media to cell culture modules, culture solution Module, thermal module and sensor assembly are preferably provided at the independent chip intracavitary.In these described schemes, the box set It may include：

It is integrated with the cabinet of fluid path drive module, gas control valve and control module；

Chip chamber；

Media to cell culture modules, culture solution module, thermal module and sensor assembly；And

Connecting tube.

In certain embodiments, independent chip intracavitary can be integrated with thermal module and sensor assembly.Therefore, herein Box set include：

It is integrated with the chip chamber of thermal module and sensor assembly；

Media to cell culture modules and culture solution module or culture chip as described herein；And

Connecting tube.

After taking the box set of the utility model, those skilled in the art can say according to what is be appended herein or according to box set Each component in box set is installed on cabinet by bright book, is made into cell culture apparatus as described herein.

Whole cells contact class material of cell culture apparatus as described herein uses disposable biological compatibility material, And pass through special disinfection and sterilization treatment.Herein, when referring to leakproofness fitting, it is intended that air tight and/or leakage can be used The means of this field routine realize air tight and/or leakage, for example, suitable binder can be used to realize.

Cultural method

Included the following steps using the method that the cell culture apparatus of this paper carries out cell culture：

(1) it provides culture solution and is previously added the culture chip of cell suspension to be cultivated；

(2) the first control switch, the second control switch, third control switch and the 4th control switch are opened, culture is formed Liquid circulation loop and cell liquid circulation loop；Control the temperature and gas composition concentration of chip chamber sealing space；

(3) after cultivating, cell is added, reagent is resuspended, open the outlet of 2 cell liquid of media to cell culture modules, close the first control System switch, third control switch and the 4th control switch, open the second control switch, the mature cell in lumen 13 are harvested Into the container of recycling.

Specifically, being preinstalled with a certain amount of cell culture fluid in culture chamber 12, lumen 13, culture chamber 14.Start Before culture, cell suspension and Porcine HGF to be cultivated are added into lumen 13, is sufficiently mixed.Culture solution module 3 from The slot 207 of media to cell culture modules 2 is inserted into, and storage chamber outlet 306, storage chamber import 307 are by hollow injection pin type bidirectional interface 311 are directed through the rubber rubber plug of culture supernatants import 205, culture supernatants outlet 206 respectively, carry out supplement culture solution. Start control module 8 simultaneously, control peristaltic pump 401 is in positive opening state, culture solution module 3 and 2 shape of media to cell culture modules At culture solution circulation loop.Stored 306 outflow of chamber outlet of culture solution in 3 storage chamber 305 of culture solution module, is cultivated through upper layer Liquid import 205 and middle level culture solution import 211 enter culture sap cavity 12, then manage interior flow path to culture solution through hollow fibre filament 209 Chamber 14 flows through culture fluid path transfer tube pipeline 213.Pass through the power drive of the peristaltic pump 401 of fluid path drive module 4, culture solution Storage chamber is flow back into via processing flow path 9a, middle level culture solution outlet 212, culture supernatants outlet 206 and storage chamber import 307 In 305.This process cycle carries out, and the fresh medium in storage chamber 305 is continuously passed through through hollow fibre filament 209 Lumen 13 participates in culture solution fluid path and exchanges.Control module 8 controls peristaltic pump 402 and is in positive opening state, lumen 13 Cell liquid import 215 is connected to processing flow path 10a with cell liquid outlet 216, and the circuit of one cycle of form ensures thin in culture Born of the same parents possess certain mobility.7 turn on sensor 701,702,703 and 704 of sensor assembly at this time monitors chip in real time respectively Temperature, oxygen concentration, gas concentration lwevel in chamber 1 and pH value.In cell cultivation process, control system 8 is according in chip chamber 1 Temperature data acquisition, control thermal module 6 semiconductor chip temperature rise or temperature drop.Control system 8 is according to oxygen in chip chamber 1 The data of gas and gas concentration lwevel acquire, and control oxygen, carbon dioxide and each gas control valve of air 503 beat ON/OFF Closed state.Control system 8 adjusts what the flow velocity of peristaltic pump 401 and 402 or prompted more renewed according to the feedback of pH value Culture solution module 3.When needing replacing culture solution module 3, incubator overhead door 103 is opened, when taking out culture chip, grab chips are drawn Existing culture solution module 3 after the slot 207 of media to cell culture modules 2 is removed, is used medicinal alcohol to wipe cell culture by mouth 308 After the medical rubber stopper surface of module carries out disinfection, new culture solution module 3 is reinstalled.After the completion of culture, cell is added Reagent is resuspended, opens the outlet of 2 cell liquid of media to cell culture modules, just the mature cell in lumen 13 can be harvested.

The present apparatus and this method culture various cells known in the art, preferably various immunocytes can be used.Cell Including but not limited to DC cells, a variety of T cells such as CIK cell, CTL cells, til cell, CAR-T cells etc..

Claims

1. a kind of media to cell culture modules, including：

The closed ontology being in communication with the outside only is exported by culture solution import, culture solution outlet, cell liquid import and cell liquid；

Be located within the body the culture sap cavity and lumen in portion, wherein the culture sap cavity include it is independent respectively with culture solution into The culture sap cavity and the culture sap cavity with culture solution outlet of mouth connection, the lumen and the cell liquid import and cell Liquid outlet；

Be placed in it is in lumen, on wall have do not allow cell by micropore hollow fibre filament；Wherein, the doughnut Silk passes through the sealing block, the both ends of hollow fibre filament respectively with the culture sap cavity of culture solution input end and culture solution outlet end Culture sap cavity be connected to.

2. media to cell culture modules as described in claim 1, which is characterized in that

The culture sap cavity is located at lumen both sides, is separated by sealing block, and hollow fibre filament passes through lumen to be connected to cell The culture sap cavity of chamber both sides；Or the culture sap cavity is located at lumen the same side, between two culture sap cavities and culture sap cavity It is separated by sealing block between lumen.

3. media to cell culture modules as claimed in claim 1 or 2, which is characterized in that

The culture sap cavity with culture solution inlet communication has culture sap cavity import, the culture sap cavity import and the culture solution Inlet communication；The culture sap cavity with culture solution outlet has the outlet of culture sap cavity, the culture sap cavity outlet and institute State culture solution outlet；

Wherein, the culture sap cavity import is located in the ontology, passes through intrinsic processing flow path and the culture solution import Connection；The culture sap cavity outlet is located in the ontology, and the media to cell culture modules further includes being located at the body interior Flow path and the optional culture fluid path transfer tube pipeline for being located at the body exterior are processed, the culture sap cavity outlet passes through described Process flow path and the optional culture fluid path transfer tube pipeline and the culture solution outlet.

4. the media to cell culture modules as described in any one of claim 1-2, which is characterized in that

There is the lumen lumen import and lumen to export, respectively with cell liquid import and cell liquid outlet；Its In, the lumen import is each provided at the body interior with lumen outlet, and the media to cell culture modules further includes being located at institute State the cell fluid path transfer tube pipeline of body exterior, wherein the lumen import is exported with lumen respectively by being located at The processing flow path and cell liquid import and cell liquid outlet, the cell liquid import for stating body interior go out with the cell liquid Pass through the cell fluid path transfer tube pipeline connection between mouthful.

5. media to cell culture modules as described in claim 1, which is characterized in that

The media to cell culture modules has culture solution flow path and cell liquid flow path；Wherein, culture solution flow path includes culture solution successively Import, the culture sap cavity of input end, inside hollow fibre filament, the culture sap cavity of outlet end and culture solution outlet；Cell liquid flow path Include cell liquid import, the hollow fibre filament external space in lumen and cell liquid outlet successively.

6. media to cell culture modules as claimed in claim 4, which is characterized in that

The media to cell culture modules has culture solution flow path and cell liquid flow path；Wherein, culture solution flow path includes culture solution successively Import, processing flow path, culture sap cavity import, the culture sap cavity of input end, inside hollow fibre filament, the culture sap cavity of outlet end, Cultivate sap cavity outlet, processing flow path, optional culture fluid path transfer tube pipeline and culture solution outlet；Cell liquid flow path includes successively Cell liquid import, processing flow path, lumen import, the hollow fibre filament external space in lumen, lumen outlet, processing stream Road, cell liquid outlet and cell fluid path transfer tube pipeline.

7. media to cell culture modules as described in claim 1, which is characterized in that the ontology of the media to cell culture modules by upper layer, Middle level and lower layer's composition, wherein：

The upper layer is equipped with culture supernatants import and culture supernatants export, and connects respectively with the culture sap cavity for being set to middle level It is logical；And it connection culture supernatants import and the processing flow path of middle level culture solution import and is connected to culture supernatants and exports and middle level The processing flow path of culture solution outlet；

The middle level is the frame structure of a middle part hollow out, which is equipped with the middle level with culture supernatants inlet communication Processing flow path, culture sap cavity between culture solution import, the import of culture sap cavity, middle level culture solution import and culture sap cavity import go out The middle level culture solution outlet, the outlet of culture sap cavity and the processing between the outlet of middle level culture solution of mouth and culture supernatants outlet Processing flow path, lumen between flow path and cell liquid import, lumen import, cell liquid import and lumen import go out Mouth, cell liquid outlet and the processing flow path between lumen outlet and cell liquid outlet；

The hollow fibre filament is placed in lumen, and both ends pass through the sealing block, is connected to respectively with culture sap cavity；With

The optional culture fluid path transfer tube pipeline being arranged in the body exterior, the culture fluid path transfer tube pipeline connection training Nutrient solution chamber is exported exports separated processing flow path with middle level culture solution；And/or

The optional cell fluid path transfer tube pipeline being arranged in the body exterior, cell fluid path transfer tube pipeline connection institute Cell liquid import is stated to export with cell liquid.

8. media to cell culture modules as claimed in claim 7, which is characterized in that

The culture fluid path transfer tube pipeline and cell fluid path transfer tube pipeline are flexible pipe for medical purpose；And/or

The upper layer is additionally provided with slot and chip pulling mouth, wherein the slot is inserted into culture solution module, the chip for positioning Pulling mouth takes out culture solution module for facilitating；And/or

Sealing block is bonded with the contact surface leakproofness in middle level, and equal leakproofness is bonded between upper layer, lower layer and middle level binding face, by The side wall of sealing block and upper layer inner wall, lower layer's inner wall and hollow-out part forms the culture sap cavity and lumen；And/or

Between the culture supernatants import and middle level culture solution import and in culture supernatants outlet and middle level culture solution Sedimeter is set between outlet, with the pollution introduced to avoid puncture medical rubber stopper；Preferably, filter sizes are 0.2 μ m。

9. media to cell culture modules as described in claim 1, which is characterized in that

The volume of the culture sap cavity is 0.2~0.5L, preferably 0.3~0.35L；And/or

The volume of the lumen is 0.4~1L, preferably 0.6~0.8L；And/or

The length of the hollow fibre filament is 130~170mm, and 20~50 μm of thickness of pipe wall, the aperture of micropore is less than 1 μm on tube wall； And/or

The ontology and sealing block are transparent biocompatible materials；And/or

Culture solution import, culture solution outlet are all made of medical rubber stopper and are sealed.

10. a kind of culture solution module, including：

11. culture solution module as claimed in claim 10, which is characterized in that

The volume of the storage chamber is 0.6~2L, preferably 0.8~1.5L；And/or

The ontology and ventilated membrane are transparent biocompatible materials；And/or

Storage chamber import, storage chamber outlet are all made of medical rubber stopper and are sealed；And/or

The storage chamber with storage liquid inlet communication has a storage chamber import, the storage chamber import by default processing pipeline with The storage liquid inlet communication；The storage chamber with storage liquid outlet has the outlet of storage sap cavity, storage chamber outlet By default processing pipeline with the storage liquid outlet；And/or

The inlet and outlet of storage chamber are equipped with sedimeter, enter in storage chamber for filtering and obstructing impurity and bacterium.

12. culture solution module as claimed in claim 10, which is characterized in that

The culture solution module is made of upper layer, middle level, lower layer and ventilated membrane, is bonded at the middle and upper levels with lower layer with middle level, middle level It combines closely between face, no leakage, is equipped with to be equipped between ventilated membrane and/or middle level and lower layer at the middle and upper levels between middle level and breathe freely Film；

Preferably, during storage liquid inlet and outlet and storage chamber inlet and outlet are all provided in middle level, when feed flow, uses hollow injection pin type Bidirectional interface, the medical rubber stopper of direct penetration cell culture module and culture solution module simultaneously.

Preferably, it is additionally provided with filling exit in the culture solution module, which is sealed using medical rubber stopper, for initially training Nutrient solution it is filling；Preferably, sedimeter is further respectively had at filling exit.

13. a kind of cell cultivation chip, the media to cell culture modules described in any one of 1-9 containing claim and claim Culture solution module described in any one of 10-12, wherein the culture solution import of the media to cell culture modules and culture solution outlet Liquid inlet communication is exported and stored respectively with the storage liquid of the culture solution module.

14. a kind of cell culture cabinet, including：Casing is integrated in fluid path drive module, the control in face in or on the cabinet Module and chip chamber for accommodating the cell cultivation chip described in claims 13, and the optional cabinet inside that is integrated in Thermal module and sensor assembly；

The control module includes：First control switch, for controlling first driving means；Second control switch, for controlling Second driving device；Third control switch, for controlling gas conveying；It is switched with the 4th optional control, for controlling temperature The heating or cooling of module.

15. the cell culture cabinet as described in claims 14, which is characterized in that

The chip chamber is independently of a container except cabinet, may be provided with air admission hole and venthole on the container, is used for It is connected to venthole with the air admission hole on cabinet, to be connected to gas module；Or

The chip chamber is a space in cabinet, the space is only equipped with by cabinet after closing cabinet air admission hole and Venthole is connected to outside.

16. the cell culture cabinet as described in claims 14, which is characterized in that

The thermal module includes semiconductor temperature-control piece and the temperature sensitive member platinum resistance that is mounted thereon, for controlling heating Or refrigeration；

The sensor assembly includes that temperature sensor, oxygen concentration sensor, gas concentration lwevel sensor and liquid pH value pass Sensor.

17. the cell culture cabinet as described in claims 14, which is characterized in that it is additionally provided with front plate on the cabinet, Upper installation display screen and control button, the work operating condition for real-time display instrument and the control to instrument operate.

18. the cell culture cabinet as described in claims 14, which is characterized in that

It is also set on cabinet with air admission hole and venthole, for being connected to the gas module；With

Also integrated 1~5 gas control valve on case is turned on and off the third control switch control by respective numbers.