CN211522186U - Filter and cell microcarrier separation device comprising same - Google Patents

Abstract

The utility model provides a filter and a cell microcarrier separation device comprising the same, wherein the cell microcarrier separation device comprises a tank body, a filter and a stirring system, wherein the filter is arranged in the tank body, and a filter membrane can be fixed on the filter; the stirring system is used for forming a vortex flow field with a vortex center positioned inside the filter in the liquid. After the stirring system is started, the microcarriers and cells outside the filter flow to the center of the vortex, when the microcarriers flow to the filter membrane on the filter, the cells with the size smaller than the aperture of the filter membrane can normally pass through, and the microcarriers with the size larger than the aperture of the filter membrane are intercepted. The utility model discloses a cell microcarrier separator utilizes vortex flow field and filter membrane separation microcarrier and cell, and operating time is fast, and the reposition of redundant personnel is efficient, is applicable to the cell sap processing of various volume scales.

Description

Filter and cell microcarrier separation device comprising same

Technical Field

The utility model belongs to the technical field of biological cell engineering, concretely relates to filter and cell microcarrier separator including this filter.

Background

Microcarrier culture is a common technique in the field of cell culture at present, and is a cell culture technique in which fine particles (about 100 μm in size) are used as a carrier and are fully mixed with a cell (about 10 μm in size) suspension, and then the cells can be attached to the surface of the carrier for culture and proliferation. The microcarrier culture has the advantages of common monolayer culture and suspension culture, and has the advantages of high cell proliferation speed, wide application range and the like. In the field of vaccine production, virus production or partial immune cell culture, microcarrier culture can be widely applied to culture of some adherent cells, culture domestication is not needed, the adherent characteristics of the cells are maintained, and a cell culture solution can be fully utilized for rapid amplification.

In microcarrier culture of adherent cells, in general, the harvesting of the cells of interest is the final critical step therein. In general, the cells are separated from the microcarriers by adding a lytic enzyme after the culture is completed and collected by centrifugation or filtration.

Cell therapy is taken as an accurate medical means, exogenous genes are mostly transfected to immune cells by adopting a lentiviral vector at present, adherent cells are used in a virus production process, the attached cells are far from a large-batch kiloliter-level production-level suspension cell bioreactor used by some conventional production enterprises, a multilayer cell factory or a microcarrier production process is mostly adopted, because the harvested products are different, a filtering and clarifying method in an antibody production process cannot be adopted, a centrifugal method can be adopted when the cells are simply collected, but when the cells and the microcarriers need to be separated after being cultured together, the simple centrifugal method cannot achieve an ideal separation effect, the complexity of the process flow is increased, in addition, the cell density of the microcarrier culture is high, and the cell density can reach 1 × 10⁷More than ml, the separation filter membrane is easily blocked when the filtration membrane is penetrated by adopting a filtration method.

Therefore, there is still a need in the art for a more efficient cell microcarrier separation device that is suitable for separating cell microcarrier mixtures of various volume scales.

SUMMERY OF THE UTILITY MODEL

The utility model provides a filter, which comprises a container frame structure and one or more blocking piece supporting pieces; the container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container and used for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece; the stopper supporting piece is arranged outside the container frame structure and fixed on the filter membrane fixing piece, and an included angle of 10-30 degrees is formed between the stopper supporting piece and a normal line of a connecting point of the stopper supporting piece and the filter membrane fixing piece.

In one or more embodiments, the filter membrane fixing member has a hollowed-out structure for fixing the filter membrane; the blocking piece supporting piece is provided with a hollow structure and used for fixing the filter membrane.

In one or more embodiments, the filter further comprises a first filter membrane secured to the filter membrane holder and a second filter membrane secured to the stopper support; the mesh size of the first filter and the second filter is between the size of the microcarrier used for cell culture and the cell size, and is used for trapping the microcarrier.

In one or more embodiments, the filter further comprises an end cap secured to the structure forming the upper opening of the container and sealing the upper opening, the end cap having two through holes.

The utility model also provides a cell microcarrier separator, include:

a tank body;

a filter disposed inside the tank; and

the stirring device is arranged in the stirring system inside the filter;

the filter comprises a container frame structure, wherein the container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container and used for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece.

In one or more embodiments, the filter is a filter according to any embodiment of the present invention.

In one or more embodiments, the separation device further comprises a liquid inlet disposed at the top or upper sidewall of the tank and a waste liquid outlet disposed at the lower sidewall or bottom of the tank.

In one or more embodiments, the cell microcarrier separation device further comprises an end cap for sealing the tank to form a closed space inside the tank, wherein the end cap is provided with two through holes for communicating the outside of the tank and the inside of the filter, one through hole is used for installing the stirring system, and the other through hole is used for discharging cells.

In one or more embodiments, the stirring system comprises a motor, a stirring shaft and a stirring paddle, wherein the stirring shaft is connected with a rotating shaft of the motor and extends into the filter.

The utility model also provides a cell microcarrier separator, include:

a tank body;

an end cap sealably connected to the tank;

a filter disposed inside the tank; and

the stirring device is arranged in the stirring system inside the filter;

wherein, a liquid inlet is arranged on the top or upper side wall of the tank body, and a waste liquid outlet is arranged on the lower side wall or bottom of the tank body;

the end cover is provided with a first through hole and a second through hole which are communicated with the outside of the tank body and the inside of the filter, the first through hole is used for installing the stirring system, and the second through hole is used for discharging cells;

the filter comprises a container frame structure and one or more stopper supports; the container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container and used for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece; the stopper supporting piece is arranged outside the container frame structure and fixed on the filter membrane fixing piece, and an included angle of 10-30 degrees is formed between the stopper supporting piece and a normal line of a connecting point of the stopper supporting piece and the filter membrane fixing piece; the filter membrane fixing part is provided with a hollow structure and is used for fixing the filter membrane; the blocking piece supporting piece is provided with a hollow structure and is used for fixing the filter membrane; the filter also comprises a first filter membrane and a second filter membrane, the first filter membrane is fixed on the filter membrane fixing piece, and the second filter membrane is fixed on the stopper supporting piece; the mesh size of the first filter and the second filter is between the size of the microcarrier used for cell culture and the cell size, and is used for trapping the microcarrier.

Drawings

FIG. 1 is a schematic structural view of the cell microcarrier separation device of the present invention.

FIG. 2 is a schematic cross-sectional view of the filter, the stirring shaft and the stirring paddle of the cell microcarrier separation device of the present invention.

The reference numerals are explained below:

1. a tank body; 2. a filter; 3. a stopper support; 4. fixing a filter membrane; 5. a motor; 6. a stirring shaft; 7. a stirring paddle; 8. a liquid inlet; 9. a cell sap outlet; 10. a waste liquid outlet; 11. an end cap; 111. a first through hole; 112. a second via.

Detailed Description

To make the features and effects of the present invention comprehensible to those skilled in the art, the general description and definitions are made with the terms and expressions mentioned in the specification and claims. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The utility model discloses a cell microcarrier separator, applicable in the cell sap processing of various volume scales, especially be applicable to the cell sap processing of the volume scale about the middle-size and small-size 1 ~ 50L.

The utility model discloses a cell microcarrier separator includes a jar body, filter and mixing system. Wherein, the filter is arranged in the tank body, and a filter membrane can be fixed on the filter; the stirring system is used for forming a vortex flow field with a vortex center positioned inside the filter in the liquid. After the stirring system is started, the microcarrier and the cells outside the filter flow to the vortex center in the filter and flow to the filter membrane on the filter, the cells with the size smaller than the pore size of the filter membrane can normally pass through the filter membrane, and the microcarrier with the size larger than the pore size of the filter membrane is intercepted, so that the microcarrier and the cells in the microcarrier cell mixed solution are separated.

In the present invention, the shape of the tank is not particularly limited, and may be, for example, a cylindrical shape or a rectangular parallelepiped shape.

The cell microcarrier separation device may also include an end cap that is sealably connected to the canister. In certain embodiments, the filter is mounted inside the canister and sealingly secured to the end cap. The end cover is provided with a first through hole which is communicated with the outside of the tank body and the inside of the filter. The stirring system comprises a motor, a stirring shaft and a stirring paddle. The motor is installed on the end cover. The stirring shaft connected with the rotating shaft of the motor extends into the filter through the first through hole on the end cover. The stirring shaft is fixed with a stirring paddle. The stirring device of the stirring system comprises a stirring shaft and a stirring paddle and is arranged inside the filter. In certain embodiments, the filter of the present invention is cylindrical.

The cell microcarrier separation device of the utility model can also comprise a liquid inlet and a liquid outlet. The liquid inlet and outlet comprises a liquid inlet, a cell sap outlet and a waste liquid outlet. The liquid inlet is positioned on the side wall or the end cover at the upper part of the tank body and is used for communicating the space outside the tank body and the space outside the filter in the tank body. The end cover is provided with a second through hole for communicating the outside of the tank body with the inside of the filter, and a cell sap outlet is formed at the second through hole. The waste liquid outlet is positioned on the side wall or the bottom of the lower part of the tank body and is used for communicating the outside of the tank body with the outside of the filter inside the tank body.

The filter of the present invention includes a container frame structure and an optional first filter membrane. The container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece connected with the structure forming the upper opening of the container and formed on the periphery of the container for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece. The container frame structure forms a structure with one end being open and the other end being closed. Herein, the open end is referred to as the top of the filter and the closed end is referred to as the bottom of the filter. The filter membrane holder is preferably cylindrical. The filter membrane fixing part is provided with a hollow structure. The first filter membrane is detachably fixed to the filter membrane fixing member, and thus, the first filter membrane may be additionally provided in the form of a consumable. The mesh size (pore size) of the first filter is set to allow cells to pass therethrough while retaining microcarriers, and is usually not more than 100 μm, and may be, for example, not more than 70 μm, 50 to 70 μm, or the like. Usually, the area of the hollow part on the filter membrane fixing part accounts for most of the total area of the filter membrane fixing part so as to fully exert the filtering function of the first filter membrane. In some embodiments, the area of the hollowed-out portion of the filter membrane fixing member accounts for more than 50% of the total area of the filter membrane fixing member, such as more than 60%, more than 70%, more than 80%, more than 90%, and the like. The fixing manner of the first filter membrane on the filter membrane fixing member is not particularly limited, for example, the filter membrane fixing member may have a slot matching with the shape and size of the first filter membrane, and the first filter membrane may be inserted into the slot and fixed on the filter membrane fixing member. In certain embodiments, the filter membrane holders have a two-layer frame structure, and the first filter membrane can be placed and clamped between the two layers of the frame. The bottom container connected to the filter membrane holder preferably has an outwardly protruding shape such as a shallow conical shape protruding outwardly, a shallow circular dish shape protruding outwardly, a spherical shape protruding outwardly, a truncated conical shape protruding outwardly, etc. to reduce liquid flow resistance and shear force to cells. Typically, the filter is sealingly secured to the end cap by structure defining an upper opening of the container. The filter may be sealingly secured to the end cap using methods conventional in the art. In some embodiments, the edge of the structure forming the upper opening of the container has a projection, the end cap has a fitting portion for fitting with the projection, and the filter is fixed to the end cap by fitting the projection into the fitting portion of the end cap; preferably, a rubber sealing ring can be arranged in the embedding part on the end cover to ensure the sealing connection between the filter and the end cover.

In certain embodiments, the filters of the present disclosure further comprise one or more stopper supports and optionally a second filter membrane. The blocking piece supporting piece has a flat-plate shape and a hollow structure. The second filter membrane is removably secured to the stopper support, and thus, the second filter membrane may be additionally provided in the form of a consumable. The stopper supporting piece and the second filter membrane form a stopper. The mesh size (pore size) of the second filter is set to allow cells to pass through the filter while retaining microcarriers, typically no more than 100 μm, and may be, for example, no more than 70 μm, 50-70 μm, etc. Usually, the area of the hollowed-out portion of the stopper support occupies a large portion of the total surface area, so as to fully exert the filtering function of the second filter membrane. In some embodiments, the area of the hollowed-out portion of the flight support can comprise more than 50% of the total surface area, such as more than 60%, more than 70%, more than 80%, more than 90%, etc. The fixing manner of the second filter membrane on the stopper support is not particularly limited, for example, the stopper support may have a slot matching the shape and size of the second filter membrane, and the second filter membrane may be inserted into the slot and fixed on the stopper support. In some embodiments, the barrier support has an elongated double frame structure, and the first filter membrane can be clamped between the two frames. The stopper supporting piece is arranged outside the container frame structure and fixed on the filter membrane fixing piece; preferably, the stopper support is disposed parallel to the axial direction of the filter membrane fixing member. The filter of the present invention may include a plurality of stopper support members. In embodiments where the filter includes a plurality of stopper supports, the plurality of stopper supports are preferably fixed to the filter membrane holder at equal intervals or symmetrically around the filter membrane holder.

In some embodiments, as shown in fig. 2, the support of the stopper is disposed to be inclined toward the rotation direction of the stirring shaft, so that the stopper can trap the microcarrier in the swirling flow field while ensuring a certain fluidity of the microcarrier, thereby avoiding blocking the second filter membrane. Preferably, the angle between the blocking piece supporting piece and the normal line at the connection point A of the blocking piece supporting piece and the filter membrane fixing piece is 10-30 degrees (shown as ^ alpha in fig. 2).

The length of the stopper support (in the axial direction of the filter) is not particularly limited, and may be the same as or smaller than the height of the filter membrane fixing member, and may be, for example, 1/5 to 4/5 or 1/2 to 4/5. The width of the stopper support is also not particularly limited and can be, for example, 1/10 to 1/2 of the filter inner diameter, or 1/5 to 1/3 of the filter inner diameter. It will be appreciated that the width-wise extension of the flight support should not touch the inner wall of the tank. When the length of the stopper supporting member is shorter than the height of the filter membrane fixing member, there is no particular limitation on the placement position of the stopper supporting member in the axial direction of the filter, for example, the stopper supporting member can be placed at the center of the outside of the filter membrane fixing member (i.e., the distance from the upper end of the stopper supporting member to the upper end of the filter membrane fixing member is the same as the distance from the lower end of the stopper supporting member to the lower end of the filter membrane fixing member), or the lower end of the stopper supporting member is flush with the lower end of.

The utility model can adopt the conventional connecting mechanism in the field, such as a shaft seal, to connect the rotating shaft of the motor with the stirring shaft. Typically, the agitator shaft is rotatably sealed through the first through bore, e.g. the junction of the agitator shaft and the first through bore may be provided with a rubber sealing ring or the like. In certain embodiments, the cell microcarrier separation device of the present invention further comprises a controller, wherein the controller is electrically connected to the motor for controlling the operation of the motor. The first through-opening is preferably located on the axis of the filter which is fixed to the end cap. Preferably, the first through opening, the axis of the stirring shaft and the axis of the filter fixed to the end cap are located on the same straight line. The stirring paddle on the stirring shaft is driven by the motor to rotate, and then liquid in the filter is stirred. The paddles may be known paddles capable of generating a vortical flow field, typically having 2 to 5 blades, and may be, for example, a propeller having 3 blades. The stirring paddle can form a vortex flow field in the liquid during rotation, so that microcarriers and cells outside the filter flow to the center of the vortex. The blades are typically characterized by large diameter, high torque, and low shear. Preferably, the edges of the blades are rounded and smooth, so that the stirring paddle cannot damage cells during rotation.

The utility model discloses in, the inlet is the inlet of the mixed liquid of microcarrier cell of treating the separation and enzyme liquid, and the cell liquid export is the liquid outlet of purpose cell, and the liquid outlet of waste liquid export is the liquid outlet of jar body waste liquid. The liquid inlet is preferably arranged on the end cover or on the side wall of the tank body close to the end cover. The waste liquid outlet is preferably arranged at the bottom of the tank body or on the side wall of the tank body close to the bottom of the tank body. The liquid inlet, the cell liquid outlet and/or the waste liquid outlet are respectively and optionally fixed with a liquid inlet pipeline, a cell liquid pipeline or a waste liquid pipeline in a sealing way, and each pipeline can be connected with a corresponding liquid storage device. In certain embodiments, the cytosol line on the cytosol outlet extends into the interior of the filter to facilitate removal of the cells of interest. The liquid inlet pipeline, the cell liquid pipeline and/or the waste liquid pipeline can be provided with a switch for controlling the smoothness and the closing of the corresponding pipelines. The liquid inlet pipeline, the cell liquid pipeline and/or the waste liquid pipeline can be provided with a driving mechanism for driving the liquid in the pipeline to flow, for example, various pumps can be arranged.

The cell microcarrier separation device of the utility model also comprises a controller which is electrically connected with one or more switches and/or driving mechanisms and is used for controlling the operation of the switches and/or the driving mechanisms.

In the utility model, the tank body, the filter membrane fixing part and the blocking part supporting piece are made of stainless steel which is easy to clean and disinfect. The materials of the first filter membrane and the second filter membrane can be polymers, metals, ceramics, composite materials and the like.

When the cell microcarrier separation device of the utility model is used for separating cells and microcarriers in microcarrier cell mixed liquor, firstly adding microcarrier cell mixed liquor to be separated into the tank body; the cells in the microcarrier cell mixture may be attached to the microcarriers or detached from the microcarriers; if the cells in the added microcarrier cell mixed solution are attached to the microcarriers, the separation enzyme solution needs to be added and fully mixed; placing the corresponding first filter membrane and the optional second filter membrane in the filter membrane fixing piece and the optional stopper supporting piece of the filter, and fixing the filter on the end cover; and covering the end cover, starting the stirring system, and driving the stirring paddle to rotate by the motor through the stirring shaft to form a vortex flow field in the liquid. At this time, both the microcarrier and the cells outside the filter flow to the center of the vortex, and when the microcarrier and the cells flow to the first filter membrane on the filter membrane fixing part of the filter, the cells can normally enter the filter through the filter membrane due to different particle sizes, and the microcarrier is blocked outside the filter.

In the embodiment that the filter comprises the stopper support member, the second filter membrane arranged on the stopper support member on the filter membrane fixing member preferentially intercepts the microcarriers in the liquid vortex, so that the liquid vortex is prevented from passing through the first filter membrane on the filter membrane fixing member too fast to cause blockage, and more cells are ensured to smoothly enter the filter through the first filter membrane on the filter membrane fixing member. In some embodiments, the stopper support is inclined at a certain angle along the rotation direction of the stirring shaft, so that the stopper can intercept not only the microcarrier in the vortex field, but also ensure that the intercepted microcarrier has certain fluidity, and avoid the loss of cells caused by the direct blockage of the microcarrier on the second filter membrane.

When the stirring system rotates for a period of time, the stable stirring speed is kept, most target cells are positioned in the center of the vortex field, and at the moment, a pipeline switch of the cell fluid outlet and a driving mechanism for driving liquid in the pipeline to flow are opened, so that the collection of target cell fluid in the center of the vortex field can be completed. During the collection process, some buffer solution can be added from the liquid inlet as appropriate, and the filter membrane on the filter is washed, so that the cell recovery rate is increased. After separation, the stirring system stops rotating, and the end cover is opened to collect a small amount of cell sap at the bottom of the filter. Opening the waste outlet allows recovery of the microcarriers trapped outside the filter.

The utility model discloses a cell microcarrier separator is applicable to the separation of microcarrier and cell of various volume scales, has following beneficial effect:

1. the cell microcarrier separation device of the utility model utilizes the vortex flow field and the filter membrane to separate microcarrier and cells, the operation time is fast, and the shunting efficiency is high;

2. in order to avoid the centrifugation process, the filter membrane of the filter is directly blocked by the microcarrier in the vortex field, the filter of the utility model is provided with the blocking piece which is arranged to be inclined at a certain angle along the rotation direction of the stirring motor, so that the microcarrier can be intercepted in the vortex field, certain fluidity of the microcarrier can be ensured, and the cell loss caused by directly blocking the filter membrane of the blocking piece is avoided;

3. the first filter membrane and the second filter membrane on the filter of the utility model are respectively detachably fixed on the filter membrane fixing part and the blocking part supporting part, so that the filter membrane fixing part and the blocking part supporting part can be conveniently removed for cleaning or replacement, and different filter membranes can be selected according to actual requirements, thereby improving the production efficiency and reducing the equipment configuration and maintenance cost;

4. use the utility model discloses a cell microcarrier separator separation accomplishes the back, can directly take out purpose cell liquid by the cell liquid export, and the simple operation has simplified the experiment operation.

The present invention will be further described with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present disclosure, and any devices and configurations similar or equivalent to those described herein may be used in the present disclosure.

Example 1

This embodiment describes a microcarrier separation device according to the present invention with reference to the drawings.

As shown in fig. 1, the separation apparatus includes a tank 1, a filter 2, a stirring system, and a liquid inlet and outlet. The tank 1 can contain a microcarrier cell mixture to be separated. The can body 1 has an end cap 11 to seal the can body 1. The material of the tank body 1 is stainless steel.

The filter 2 comprises a stopper support 3 and a vessel frame structure, optionally further comprising a first filter membrane and a second filter membrane. The container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece 4 which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container for fixing a filter membrane, and a bottom container connected with the fixing piece. The stopper support 3 is made of stainless steel. The stopper support 3 has a strip-shaped double-layer frame structure, and a second filter membrane can be placed in the middle. The stopper supporting piece 3 and the second filter membrane form a stopper. The material of the filter membrane fixing part 4 is stainless steel. The filter membrane holder 4 is cylindrical. The bottom container connected to the filter membrane holder 4 is in the shape of a shallow cone protruding downwards. The filter membrane fixing part 4 has a double-layer frame structure, and a first filter membrane can be placed in the middle. The first filter membrane is fixed on the filter membrane fixing part 4. The first filter membrane and the second filter membrane are made of polymers. The mesh size of the first and second filter is set to allow cells to pass through the filter and retain microcarriers, typically no more than 100 μm. The stopper supporting piece 3 is fixed on the filter membrane fixing piece 4, and the length direction of the stopper supporting piece 3 is parallel to the axial direction of the filter membrane fixing piece 4. The filter 2 is positioned in the tank body 1, and the open end of the filter 2 is hermetically fixed on the end cover 11.

The stirring system comprises a motor 5, a stirring shaft 6 and a stirring paddle 7. The motor 5 is fixed on the end cover 11. The stirring shaft 6 is connected with the rotating shaft of the motor 5. The stirring shaft 6 passes through the first through hole 111 of the end cover 11 and enters the inside of the filter 2. The stirring paddle 7 is fixed on the stirring shaft 6. The stirring paddle 7 is driven by the motor 5 to rotate, so that the liquid in the filter 2 is stirred, and a vortex flow field is formed in the liquid.

As shown in fig. 2, flight support 3 is disposed so as to be inclined toward the rotational direction of stirring shaft 6. The angle between the blocking piece supporting piece 3 and the normal line at the connecting point A of the blocking piece supporting piece 3 and the filter membrane fixing piece 4 is 10-30 degrees (namely ^ alpha in fig. 2). Six baffle supports 3 are fixed on the filter membrane fixing part 4 in a surrounding way at equal intervals.

The liquid inlet and outlet comprises a liquid inlet 8, a cell liquid outlet 9 and a waste liquid outlet 10. The liquid inlet 8 is a liquid inlet for the microcarrier cell mixed liquid and the enzyme liquid to be separated. The cell fluid outlet 9 is a fluid outlet of the target cell. The waste liquid outlet 10 is a liquid outlet of waste liquid in the tank body 1. The liquid inlet 8 is positioned on the end cover 11 and is used for communicating the outside of the tank body 1 with the outside part of the filter 2 inside the tank body 1. The cell sap outlet 9 passes through the second through hole 112 of the end cap 11 for communicating the outside of the tank 1 and the inside of the filter 2. The waste liquid outlet 10 is located below the side wall of the tank 1 for communicating the outside of the tank 1 with the outside of the filter 2 inside the tank 1. A liquid inlet pipeline, a cell liquid pipeline and a waste liquid pipeline are respectively fixed on the liquid inlet 8, the cell liquid outlet 9 and the waste liquid outlet 10 in a sealing way, and each pipeline is provided with a switch for controlling the smoothness and the closing of the pipeline.

Example 2

This example describes the procedure of microcarrier and cell separation using the microcarrier separation device of example 1 with reference to the accompanying drawings.

When the separating device works, firstly, the microcarrier cell mixed liquid to be separated and the separated enzyme liquid are added from the liquid inlet 8 and are fully mixed. Filter membranes (with the aperture of 50-70 μm) are placed in the filter membrane fixing piece 4 and the stopper supporting piece 3 of the filter 2, and the filter 2 is fixed on the end cover 11. The end cover 11 is covered, the stirring system is started, the motor 5 drives the stirring paddle 7 to rotate rapidly through the stirring shaft 6, and a vortex flow field is formed in the liquid. At this time, both the microcarriers and cells outside the filter 2 flow toward the center of the vortex, and when they flow to the first filter membrane on the filter membrane holder 4 of the filter 2, the cells can normally pass through and enter the inside of the filter 2 due to the difference in particle size, while the microcarriers are blocked from the outside.

Meanwhile, the second filter membrane arranged on the stopper supporting piece 3 on the filter membrane fixing piece 4 intercepts the microcarriers in the liquid vortex preferentially, so that the liquid vortex is prevented from blocking due to the fact that the liquid vortex passes through the first filter membrane on the filter membrane fixing piece 4 too fast, and more cells are guaranteed to enter the filter 2 through the first filter membrane smoothly.

Keep off piece support piece 3 and set up to the rotation direction slope of orientation (mixing) shaft 6 for keep off piece support piece 3 not only can intercept the microcarrier in the vortex field, can also guarantee that the microcarrier of intercepting has certain mobility, avoids directly after the stifled second filter membrane that keeps off on piece support piece 3 of microcarrier, arouses the loss of cell.

When the stirring system rotates for a period of time, the stable stirring speed is kept, most target cells are positioned in the center of the vortex field, and at the moment, the pipeline switch of the cell sap outlet 9 and the driving mechanism for driving liquid in the pipeline to flow are opened, so that the collection of the target cell sap in the center of the vortex field can be completed. In certain embodiments, some buffer may be added from loading port 8 during collection as appropriate to wash the filter membrane on filter 2 and increase cell recovery. After the separation is completed, the stirring system stops rotating, and the end cover 11 is opened to collect a small amount of cell sap at the bottom of the filter 2. Opening the waste outlet 10 allows recovery of the microcarriers trapped outside the filter 2.

Claims (10)

1. A filter, comprising a container frame structure and one or more stopper supports; the container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container and used for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece; the stopper supporting piece is arranged outside the container frame structure and fixed on the filter membrane fixing piece, and an included angle of 10-30 degrees is formed between the stopper supporting piece and a normal line of a connecting point of the stopper supporting piece and the filter membrane fixing piece.

2. The filter of claim 1, wherein the filter membrane fixing member has a hollowed-out structure for fixing the filter membrane; the blocking piece supporting piece is provided with a hollow structure and used for fixing the filter membrane.

3. The filter of claim 1, further comprising a first filter membrane secured to the filter membrane securing member and a second filter membrane secured to the barrier support member; the mesh size of the first filter and the second filter is between the size of the microcarrier used for cell culture and the cell size, and is used for trapping the microcarrier.

4. The filter of claim 1 further comprising an end cap secured to the structure forming the upper opening of the container and sealing the upper opening, the end cap having two through holes.

5. A cell microcarrier separation device, comprising:

a tank body;

a filter disposed inside the tank; and

the stirring device is arranged in the stirring system inside the filter;

the filter comprises a container frame structure, wherein the container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container and used for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece.

6. The cell microcarrier separation device of claim 5, wherein the filter is according to any one of claims 1-4.

7. The cell microcarrier separation device of claim 5, further comprising a liquid inlet disposed on the top or upper sidewall of the tank and a waste liquid outlet disposed on the lower sidewall or bottom of the tank.

8. The cell microcarrier separation device of claim 5, further comprising an end cap for sealing the tank to form a closed space inside the tank, wherein the end cap has two through holes communicating the outside of the tank and the inside of the filter, wherein one through hole is used for installing the agitation system, and the other through hole is used for discharging cells.

9. The cell microcarrier separation device of claim 5, wherein the stirring system comprises a motor, a stirring shaft and a stirring paddle, and the stirring shaft is connected with the rotating shaft of the stirring paddle and the motor and extends into the filter.

10. A cell microcarrier separation device, comprising:

a tank body;

an end cap sealably connected to the tank;

a filter disposed inside the tank; and

the stirring device is arranged in the stirring system inside the filter;

wherein, a liquid inlet is arranged on the top or upper side wall of the tank body, and a waste liquid outlet is arranged on the lower side wall or bottom of the tank body;

the end cover is provided with a first through hole and a second through hole which are communicated with the outside of the tank body and the inside of the filter, the first through hole is used for installing the stirring system, and the second through hole is used for discharging cells;

the filter comprises a container frame structure and one or more stopper supports; the container frame structure comprises a structure forming an upper opening of the container, a filter membrane fixing piece which is connected with the structure forming the upper opening of the container and is formed on the periphery of the container and used for fixing a filter membrane, and a bottom container connected with the filter membrane fixing piece; the stopper supporting piece is arranged outside the container frame structure and fixed on the filter membrane fixing piece, and an included angle of 10-30 degrees is formed between the stopper supporting piece and a normal line of a connecting point of the stopper supporting piece and the filter membrane fixing piece; the filter membrane fixing part is provided with a hollow structure and is used for fixing the filter membrane; the blocking piece supporting piece is provided with a hollow structure and is used for fixing the filter membrane; the filter also comprises a first filter membrane and a second filter membrane, the first filter membrane is fixed on the filter membrane fixing piece, and the second filter membrane is fixed on the stopper supporting piece; the mesh size of the first filter and the second filter is between the size of the microcarrier used for cell culture and the cell size, and is used for trapping the microcarrier.

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