KR101091052B1 - Device for centrifugal separation and method using the same - Google Patents

Device for centrifugal separation and method using the same Download PDF

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KR101091052B1
KR101091052B1 KR1020090015016A KR20090015016A KR101091052B1 KR 101091052 B1 KR101091052 B1 KR 101091052B1 KR 1020090015016 A KR1020090015016 A KR 1020090015016A KR 20090015016 A KR20090015016 A KR 20090015016A KR 101091052 B1 KR101091052 B1 KR 101091052B1
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South Korea
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separation space
separation
tissue
bag
space
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KR1020090015016A
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Korean (ko)
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KR20100095952A (en
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정광회
이광형
최행석
신준호
고정재
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차의과학대학교 산학협력단
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/505—Containers for the purpose of retaining a material to be analysed, e.g. test tubes flexible containers not provided for above
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0631—Purification arrangements, e.g. solid phase extraction [SPE]
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0684—Venting, avoiding backpressure, avoid gas bubbles
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/12—Specific details about materials
    • B01L2300/123—Flexible; Elastomeric
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0409—Moving fluids with specific forces or mechanical means specific forces centrifugal forces
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5021—Test tubes specially adapted for centrifugation purposes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
    • C12M47/04—Cell isolation or sorting

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Hematology (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Centrifugal Separators (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

Disclosed are a centrifugal separator and a centrifugal separation method for separating target cells from tissues extracted from living organisms using centrifugation. The separator for centrifugation includes an upper separating bag of a flexible material, and a lower separating bag, and includes a partition member for selectively opening and closing a passage connecting the separating spaces inside the two separating bags. The upper portion of the upper separation bag includes a tissue inlet for injecting tissue, a treatment solution inlet for injecting a treatment solution reacting with the tissue, and a solution outlet for discharging residual material in the upper separation space after centrifugation. In the lower portion of the lower separation bag is formed a target outlet for discharging the target cells collected in the lower separation space after centrifugation. Since the injection of tissue and treatment solution and the discharge of residual material and target cells are made through different passages, they can be prevented from being mixed with each other and through a simple manufacturing process through fusion using a material such as plastic. Manufacturing is possible. In addition, there is an advantage that can easily increase the amount of the target cells to be obtained by connecting a plurality of centrifuge separator.

Tissue inlet, treatment solution inlet, solution outlet, separator for centrifugation

Description

Centrifuge Separator and Centrifugal Separation {DEVICE FOR CENTRIFUGAL SEPARATION AND METHOD USING THE SAME}

The present invention relates to a centrifugal separator and a centrifugal separation method for separating target cells from tissues extracted from living organisms using centrifugation.

Stem cells of adipose tissue can be used for the purpose of regenerating various damaged tissues in the human body, and in particular, they can be differentiated into hematopoietic cells, nerve cells, skin reconstructions, chondrocytes, cardiomyocytes, and blood vessel cells. Can be used.

In particular, the usefulness of adipose stem cells may be of great practical value compared to stem cells derived from other organs. Specifically, the adipose stem cells can be separated from the excess adipose tissue in the human body, can be obtained only with a relatively simple treatment. However, when stem cells are separated from bone marrow, umbilical cord blood, peripheral blood, and embryos, it is necessary to overcome complex problems such as ethical issues, pain of the subjects to be collected, contamination by external factors such as bacteria and viruses during the process.

However, the enzyme treatment method used to separate fat stem cells is easily exposed to external factors such as bacteria and viruses in the process and process of transferring adipose tissue collected by liposuction into a separate container. After enzymatic treatment, centrifugation for extracting only the target cells is performed again in a separate space, so that exposure to external factors is frequent during enzymatic treatment and centrifugation.

The present invention provides a centrifugal separation device and a centrifugal separation method capable of performing a plurality of processing steps in one container to obtain a desired target cell from a tissue.

In the present invention, the injection of tissues required in the process of obtaining the target cells from the tissues, the injection of the treatment solution, the discharge of residual material generated after the reaction of the treatment solution and the tissue, and the discharge of the desired target cells are performed through different passages. It provides a centrifugal separator and centrifugal separation method that can be prevented from being mixed with each other in detail.

The present invention provides a centrifugal separator that can be manufactured through a simple manufacturing process such as fusion using a flexible material such as plastic.

The present invention can be used in a single use to reduce the manufacturing cost, thereby providing a centrifugal separation device that can extract a more pure target cells.

The present invention provides a separator for centrifugation that can easily increase the amount of the target cell to be obtained by easily increasing the capacity of the separator.

The present invention provides a separator for centrifugation having a simple structure by designing the space in the separator to be treated so that the tissue to be treated, the treatment solution, and the space to recover the treated tissue are not provided separately.

According to an exemplary embodiment of the present invention for achieving the above object of the present invention, the centrifugal separator for separating the target cells from the tissue extracted from the organism using centrifugation, the upper separation bag, the lower A separating bag, and a partition member.

The upper separation bag forms an upper separation space using a flexible material, a tissue inlet for injecting tissue into the upper separation space, a treatment solution inlet for injecting treatment solution separately from the tissue inlet, and a centrifugal separation chamber in the upper separation space. Solution outlet for discharging residual material.

The lower separating bag is also formed integrally with the upper separating bag using a flexible material, forms a lower separating space connected to the upper separating space, and includes an objective outlet for discharging the target cells from the lower separating space.

The partition member selectively opens and closes a passage connecting the upper separation space and the lower separation space. Preferably it may be located between the upper separation space and the lower separation space.

The width of the upper separation space of the upper separation bag described above may be gradually reduced toward the lower separation bag from a specific point, whereas the width of the lower separation space of the lower separation bag along the longitudinal direction of the lower separation bag may be maintained to be the same. Can be.

In addition, the upper separation bag may include a valve member for selectively opening and closing the tissue inlet, the treatment solution inlet, and the solution outlet, and likewise, the lower separation bag may also include a valve member for selectively opening and closing the target outlet. The valve member may use a forceps, a valve, a zipper, or the like.

In addition, the lower separation bag can be separated from the upper separation bag in a closed state by using the partition member, the partition member may be used tongs, valves, zippers and the like.

In addition, according to another exemplary embodiment of the present invention, comprising a film separation bag formed by fusing at least one or more resin films formed using a synthetic resin, the inside of the film separation bag provides an upper separation space and a lower separation space The upper portion of the upper separation space is provided with a tissue inlet for injecting tissue, a treatment solution inlet for injecting treatment solution separately from the tissue inlet, and a solution outlet for discharging residual material in the upper separation space after centrifugation. A lower portion of the lower separation space is formed with a target discharge port for discharging the target cells collected in the lower separation space, the compartment for selectively opening and closing the passage connecting the upper separation space and the lower separation space between the upper separation space and the lower separation space. The member is provided.

The film separation bag formed by fusion of at least one resin film formed by using a flexible synthetic resin through pressure, heat, high frequency, chemical reaction, or the like, may be formed by injecting a tissue inlet, a treatment solution, or the like. The edges of the film separator bag, excluding the inlet, the solution outlet, and the target outlet, are sealed by fusion, and in particular, the width of the upper separation space may be fused and sealed such that the width of the upper separation space gradually decreases, such as a funnel toward the lower separation space, while The width of the separation space can be fused so as to remain the same. In this case, when using a single resin film can be folded and fused to overlap the resin film of different parts, it may be fused by overlapping a plurality of resin films.

In addition, it may include a tissue inlet formed in the film separation bag above the upper separation space, the treatment solution inlet, and a valve member for selectively opening and closing the solution outlet, and likewise the target outlet formed in the film separation bag below the lower upper separation space It may include a valve member for selectively opening and closing. The valve member can use a forceps, a valve or the like.

In addition, the lower separation space may be separated from the upper separation bag in a closed state by using the partition member, the partition member may use a forceps or a valve.

At this time, an incision line is formed in the film separation bag along the outer space of the lower separation space, and the lower separation space is easily closed from the film separation bag in a closed state, and after centrifuging the tissue, the target cell is separated from the lower separation bag. It can also be stored separately in the state it is stored in.

In addition, as the material of the upper separation bag, the lower separation bag, and the film separation bag of the above-described centrifugal separator, a synthetic resin of a flexible material may be used, and in particular, there is no absorbency but elasticity such as polyvinyl chloride. You can use a material that is also resistant to chemicals.

However, when the separator for centrifugation according to the present invention is used for the purpose of culturing the target cells extracted from the tissue as well as the centrifugation, the material of the upper separation bag, the lower separation bag, and the film separation bag, in addition to the above-described characteristics It is also possible to use materials through which certain gases can permeate.

For example, a polymer separation membrane that separates gas may use an oxygen enrichment membrane that cannot permeate liquid but only permeate oxygen when cultivating. This may utilize polymers of dimethylsiloxane, and polycarbonate, as is already known.

In addition, according to another exemplary embodiment of the present invention, the centrifugal separation assembly that can be used by combining two or more of the above-described centrifugal separation apparatus, the centrifugal separation apparatus of one centrifugal separation apparatus is different It is connected to the tissue inlet of the separation device for separation.

In addition, according to another exemplary embodiment of the present invention, the centrifugation method for separating the target cells from the tissue extracted from the organism using the centrifugation, forming an upper separation space using a flexible material (flexible) And a tissue inlet for injecting tissue into the upper separation space, at least one treatment solution inlet for injecting treatment solution separately from the tissue inlet, and a solution outlet for discharging the remaining material in the upper separation space after centrifugation. Upper separating bag; A lower separating bag which is formed integrally with the upper separating bag using a flexible material, forms a lower separating space connected to the upper separating space, and has a target discharge port for discharging the target cells from the lower separating space; And a partition member for selectively opening and closing a passage connecting the upper separation space and the lower separation space; Injecting tissue into the upper separation space through the tissue injection port; Injecting a treatment solution reacting with the tissue into the upper separation space through any one of the treatment solution inlets; Placing and centrifuging the upper separating bag and the lower separating bag in the order of centrifugal rotation; Separating the upper separation space and the lower separation space by using a partition member; And discharging the residual material through the solution outlet.

In addition, after the step of discharging the residual material to the solution outlet, it may further comprise the step of reprocessing the tissue treated by the treatment solution at least once using a different treatment solution separate from the treatment solution, Reprocessing may include connecting the upper separation space and the lower separation space by using a partition member; Injecting another treatment solution through any one of the treatment solution inlets; Placing and centrifuging the upper separating bag and the lower separating bag in the order of centrifugal rotation; Separating the upper separation space and the lower separation space by using a partition member; And discharging the residual material through the solution outlet.

For reference, the treatment solution herein includes not only enzymes or chemicals that react with the tissues, but also chemicals that are irrelevant to the tissues, such as washing liquids injected into the centrifugal separator and washed therein. Can be.

The centrifugal separator and centrifugal separation method of the present invention can be carried out in one container several times to obtain a desired target cell from the tissue.

Centrifugal separation apparatus and centrifugation method of the present invention is the injection of the necessary tissue in the process of obtaining the target cells from the tissue, the injection of the treatment solution, the discharge of residual substances generated after the reaction of the treatment solution and the tissue, and the desired target cells Through the discharge of the through different passages to prevent the above-mentioned materials are mixed with each other, the material to be injected by the user and the pre-injected material can enter the internal space at the same time to prevent the unwanted reaction occurs. .

The centrifuge separator of the present invention provides a centrifuge separator that can be manufactured through a simple manufacturing process such as fusion using a flexible material such as plastic. Therefore, it is possible to use a single use with a reduction in the manufacturing cost, thereby allowing the extraction of more pure target cells.

When using adipose cells to obtain a target cell such as adipose stem cells, there may be a problem that the pretreated material remains because the washing inside the separation space for repetitive treatment does not wash well in the case of fat cells. Centrifugal separator of the present invention can be used only once can solve the above problems.

The centrifugal separator of the present invention can be used by connecting a plurality of methods by connecting the target outlet of one centrifugal separator and the tissue inlet of the other centrifugal separator to easily increase the amount of the target cells to be obtained. have.

Hereinafter, preferred embodiments of the present invention will be described in detail with reference to the accompanying drawings, but the present invention is not limited or limited by the embodiments. For reference, in the present description, the same numbers refer to substantially the same elements, and may be described by quoting contents described in other drawings under such a rule, and the contents repeated or deemed apparent to those skilled in the art may be omitted.

Example 1

1 is a view showing a separator for centrifugation according to a first embodiment of the present invention.

Referring to FIG. 1, the centrifugal separator 100 for separating target cells from tissues extracted from living organisms using centrifugation may include an upper separation bag 110, a lower separation bag 120, and a partition member ( 130).

The upper separation bag 110 forms an upper separation space 111 using a flexible material, and separates the treatment solution from the tissue injection port 112 and the tissue injection port 112 for injecting the tissue into the upper separation space 111. The first treatment solution injection port 113, the second treatment solution injection port 114 for injection, and a solution outlet 115 for discharging the residual material in the upper separation space 101 after centrifugation. For reference, two treatment solution inlets 113 and 114 for injecting the treatment solution are provided in this embodiment, but may be further formed by the amount of treatment solution desired by the user.

In some cases, a plurality of treatment solutions may be injected through one treatment solution inlet, but the pre-injected treatment solution remains in the treatment solution inlet and then injected into the upper separation space 101 together with the treatment solution to be injected. It is preferable that the number is determined according to the number of types of processing solution to be used so as to prevent it.

The lower separating bag 120 is also formed integrally with the upper separating bag 110 using a flexible material, and forms a lower separating space 121 connected to the upper separating space 111, and from the lower separating space 121. And a target outlet 122 for discharging the target cell.

The partition member 130 is positioned between the upper separation space 111 and the lower separation space 121 to selectively open and close a passage connecting the upper separation space 111 and the lower separation space 121.

The upper separation bag 110 is formed in a funnel shape in which the width of the upper separation bag 110 gradually decreases toward the lower separation bag 120, so that the target cells are easily separated from the lower separation bag 120 during centrifugation. The width of the lower separation space 121 of the lower separation bag 120 is smaller than the width of the upper separation space 111 of the upper separation bag 110, and the lower separation bag 120. The width of the lower separation space 121 along the longitudinal direction of) is formed to be the same.

In addition, the upper separation bag 110 is a valve member 101 for selectively opening and closing the tissue inlet 112, the first treatment solution inlet 113, the second treatment solution inlet 114, and the solution outlet (115) In addition, the lower separation bag 120 also includes a valve member 101 for selectively opening and closing the target outlet 122.

In the present embodiment, the valve member 10 may be provided separately from tongs, valves, and zippers, but the upper and lower separating bags 110 and 120 of the present embodiment are formed of a plastic material having flexibility, so that the structure is actually organized. The inlet 112, the first treatment solution inlet 113, the second treatment solution inlet 114, the solution outlet 115, and the target outlet 122 are all provided in a blocked state, and a cannula, and a needle. Sharp and thin tubes can be used to inject or discharge the desired material. That is, when the valve member 10 is made of a synthetic film, the valve member 10 may be integrally formed with the upper and lower separation bags 110 and 120, and the centrifugal separation system may be immediately implemented by simply connecting the pipes. It is also easy to manufacture.

In addition, the upper and lower separating bags (110, 120) has a sealing property with the outside, using a plastic material having an elasticity, such as polyvinyl chloride (polyvinyl chloride) is not absorbent but elastic and high resistance to chemicals Can be used.

However, when the centrifugal separator 100 according to the present invention is used for the purpose of culturing the target cells extracted from the tissue as well as centrifugal separation, it is also possible to use a material that can penetrate a specific gas in addition to the above characteristics. For example, a liquid may not permeate, but an oxygen enrichment membrane prepared using a polymer of dimethylsiloxane and polycarbonate to permeate only oxygen may be used.

Thus, the centrifugal separator 100 of the present invention is the injection of the tissue required in the process of obtaining the target cells from the tissue, the injection of the treatment solution, the discharge of residual substances generated after the reaction of the treatment solution and the tissue, and the desired purpose The discharge of the cells through the different passages to prevent the above-mentioned materials are mixed with each other, by using the valve member 10, or by using a sharp tool such as cannula, external dust or viruses, It is thoroughly blocked from contaminants such as bacteria.

In addition, the lower separation bag 120 may be separated from the upper separation bag 110 in a sealed state by using the partition member 130, and the partition member 130 may use a forceps or a valve.

Therefore, after centrifuging the tissue using the apparatus 100, the target cells may be stored separately in the state of being stored in the lower separation bag 120.

Example  2

The centrifugal separator according to the second embodiment of the present invention is substantially the same as the centrifugal separator described in the first embodiment. Therefore, the description of the centrifugal separator in the second embodiment may refer to the description and the drawings of the centrifugal separator of the first embodiment, and repeated descriptions may be omitted. In addition, in the second embodiment of the present invention, for the convenience of description, the same components will be described using the same reference numerals.

However, in the second embodiment of the present invention will be described in more detail with respect to the manufacturing method of the centrifugal separator.

2 is a plan view of a separator for centrifugation according to a second embodiment of the present invention.

In order to manufacture a centrifuge separation apparatus 200 for separating target cells from tissues extracted from living organisms using centrifugation, first, resin films using two sheets of synthetic resins are disposed to face each other. Two resin films formed using a flexible synthetic resin may be fused through pressure, heating, high frequency, and chemical reaction to form a film separation bag 20.

In addition, the above-mentioned resin film has an airtightness with the outside, using a plastic material having elasticity, there is no water absorption, such as polyvinyl chloride (polyvinyl chloride) can be used a material having a high elasticity and resistance to chemicals.

However, when the centrifugal separator 200 according to the present invention is used for the purpose of culturing the target cells extracted from the organization as well as centrifugal separation, it is also possible to use a material that can transmit a specific gas in addition to the above characteristics. have.

Referring to FIG. 2, the inside of the film separation bag 20 provides an upper separation space 211 and a lower separation space 221, and a tissue injection hole 212 for injecting tissue into the upper separation space 211. ), The first and second treatment solution inlets 213 and 214 for injecting treatment solution separately from the tissue inlet 212, and the solution outlet 215 for discharging the residual material in the upper separation space 211 after centrifugation. ) Is provided, and a lower portion of the lower separation space 221 is formed with a target discharge port 222 for discharging the target cells collected in the lower separation space 221, and the upper separation space 211 and the lower separation space 221. The partition member 230 for selectively opening and closing the passage connecting the upper separation space 211 and the lower separation space 221 is provided between the ().

The above-described two films 10 are fused to seal the upper separation space 211 and the lower separation space 221 with the outside, and the partition member 230 is the upper separation space 211 and the lower separation space 221. You can use forceps placed between them to separate them.

In addition, when the film separation bag 20 is fused through the above-described method, the tissue inlet 212, the solution outlet 215, the first and second treatment solution inlets 213 and 214, and the target outlet ( The edge of the film separation bag 20 except for 222 is sealed by the fusion portion A formed by fusion, and in particular, the width of the upper separation space 211 toward the lower separation space 221 is fused to decrease gradually. On the other hand, the width of the lower separation space 221 may be fused so that the width of the lower separation space 221 remains the same. In some cases, the material collected in the lower separation space 221 may be circulated around the target outlet 222. Dense, such that the lower separation space 221 is also fused in a U-shaped inclined gradually smoothly in a direction opposite the upper separation space 211 so as to facilitate the acquisition of the material through the target outlet 222. , Even in the form of an inclined V-shaped Can.

Thereafter, as illustrated in FIG. 2, a valve for selectively opening and closing the tissue inlet 212, the first and second treatment solution inlets 213 and 214, the solution outlet 215, and the target outlet 222. Valve members 201 such as tongs may be disposed.

However, since the film separation bag 20 according to the present invention is formed of a plastic material having a predetermined elasticity, the tissue inlet 212, the first treatment solution inlet 213, the second treatment solution inlet 214, Both the solution outlet 215 and the target outlet 222 may be fused and provided in a clogged state, and a desired material may be injected or discharged using a sharp thin tube such as a cannula and a needle.

In addition, the incision line (L) is formed in the film separation bag (20) along the periphery of the lower separation space (221), after centrifuging the tissue using the centrifugal separation device 200 of the present embodiment, The lower separation space 221 may be cut from the film separation bag 20 in a sealed state. In particular, the boundary between the upper separation space 211 and the lower separation space 221 may be separated from the film separation bag 20 in a blocked state using the partition member 230.

Centrifugal separator 200 of the present embodiment by using a flexible material such as plastic by providing a centrifugal separator 200 that can be manufactured through a simple manufacturing process such as fusion, it is possible to reduce the manufacturing cost To this end, it is possible to use the centrifugal separator 200 for single use without burden. In addition, since it is not necessary to reuse, a more pure target cell can be extracted.

Example 3

Hereinafter, a centrifugal separation assembly that can be used for centrifugation by connecting at least two or more centrifugal separation apparatuses of the first and second embodiments described above, and the centrifugal separation according to the third embodiment of the present invention. The components of the separation assembly for separation are substantially the same as the centrifugal separation device described in the first and second embodiments. Therefore, the third embodiment will be described mainly by the connection structure between the centrifugal separators described in the first and second embodiments, and the description of the other components is the centrifugation of the first and second embodiments. Reference may be made to the description and drawings of the separation device for the device, and repeated descriptions thereof will be omitted. In addition, the same reference numerals as those of the first and second embodiments are used for components of the separation assembly for centrifugation for convenience of description.

3 is a plan view of a separation assembly for centrifugation according to a third embodiment of the present invention.

Referring to FIG. 3, two centrifugal separators 200 and 200 ′ shown in FIG. 2 are arranged side by side to form a centrifugal separation assembly 300.

At this time, the target outlet 222 of one centrifugal separator 200 is connected to the tissue inlet 212 'of the other centrifugal separator 200'. In this case, the on / off valve 201 mounted on the target outlet 222 and the on / off valve 201 'mounted on the tissue injection port 212' are always open.

In addition, between the target outlet 222 and the tissue inlet 212 'is connected to the connection pipe 30 is connected to connect the inside of the two centrifugal separator (200, 200').

Therefore, the tissue to be treated is injected through the tissue injection port 201 of the centrifugal separator 200, reacts with the treatment solution, and the target cells obtained by centrifugation are the purpose of the centrifugal separator 200 '. It can be obtained through the outlet 222 ′.

As described above, by using a plurality of centrifugal separation apparatus of the present invention by connecting, it is possible to easily increase the amount of the target cells to be obtained.

Of course, the above-described two centrifugal separators 200 and 200 'or more centrifugal separators are used in a row when used in a centrifuge.

Example 4

The centrifugal separation method according to the fourth embodiment of the present invention can be implemented using the centrifugal separation device and the centrifugal separation assembly described in the first to third embodiments, and thus, the centrifugal separation method of the present embodiment. For the description of the apparatus used, reference may be made to the contents described in the first to third embodiments, and the repeated content is omitted. In addition, for the convenience of explanation, the centrifugal separation device illustrated in the fourth embodiment will be described taking the separation device described in the second embodiment as an example. Incidentally, the same reference numerals as those in the second embodiment are used for the components of the separator.

4 is a flowchart illustrating a centrifugal separation method according to a fourth embodiment of the present invention.

Referring to FIG. 4, in the centrifugal separation method of the present embodiment, a step of preparing a centrifugal separation apparatus 200 (S410) and injecting tissue into the upper separation space 211 through a tissue injection port 201 (S420). ), Injecting the treatment solution reacting with the tissue into the upper separation space through any one of the first and second treatment solution inlets 213 and 214 (S430), the upper separation bag from the rotary shaft of the centrifugation, and the lower separation Arranging and centrifuging in white order (S440); Isolating the upper separation space and the lower separation space using the partition member (S450), and discharging the residual material through the solution outlet (S460).

This will be described using the process of separating adipose stem cells from adipose tissue as an example.

First, the aspirated adipose tissue is injected into the upper separation space 211 using a tissue injection hole 201.

Thereafter, in order to inject the enzyme treatment and washing solution for the adipose tissue, the centrifugal separator 200 is placed in a shake incubator, and the enzyme (collagenase type I) is injected through the first treatment solution inlet 213. And reacted at 37 ° C. for 2 hours.

Then, the film separation bag 20 is placed so as to be placed in the order of the upper separation space 211, and the lower separation space 221 in the rotation axis of the centrifugation, and centrifuged at 300xg for 10 minutes to recover the fat stem cells. Most of the recovered fat stem cells are collected in the lower separation space 221.

As described above, fat stem cells may be recovered and used, but if a high purity fat stem cells are desired, the following reprocessing process (S470) may be performed.

Hereinafter, the reprocessing process will be described in detail.

First, the lower separation space 221 is separated from the upper separation space 211 by using the partition member 230, and the recovered cells are prevented from moving, and then, in the upper separation space 211 through the solution outlet 215. Remove residual material, ie supernatant.

Thereafter, the washing solution is introduced through the second treatment solution inlet 214, and the partition member 230 is opened again and suspended in the stirrer for a predetermined time.

After that, centrifuge again at 300xg for 10 minutes.

Thereafter, the lower separation space 221 is separated from the upper separation space 211 using the partition member 230, and the supernatant containing the external factor added during the enzymatic treatment through the solution outlet 215 is removed.

If another reprocessing is desired, the above process can be repeated.

Specifically, the partition member 230 is opened to communicate the upper separation space 211 and the lower separation space 221, and dissolve red blood cells through the first treatment solution inlet 213 or the second treatment solution inlet 214. Add the liquid and suspend in the stirrer.

After that, centrifuge at 300xg for 10 minutes.

Thereafter, the lower separation space 221 is separated from the upper separation space 211 using the partition member 230, and the solution outlet 215 is removed from the supernatant.

Thereafter, if necessary, the enzyme reaction, the addition and stirring of the washing solution, the centrifugation and the supernatant removal process may be further repeated.

As described above, it is possible to obtain a high-purity fat stem cells collected in the lower separation space 221 through repeated enzymatic reaction, washing, and centrifugation.

At this time, the high-purity fat stem cells can be discharged to the outside through the target outlet 222, or recovered by a syringe or the like, and can be used according to the purpose. As described above, only the lower separation space 221 is separated and separately. After storage, it can be used as needed.

As described above, although described with reference to a preferred embodiment of the present invention, those skilled in the art will be variously modified and changed without departing from the spirit and scope of the invention described in the claims below I can understand that you can.

1 is a view showing a separator for centrifugation according to a first embodiment of the present invention.

2 is a plan view of a separator for centrifugation according to a second embodiment of the present invention.

3 is a plan view of a separation assembly for centrifugation according to a third embodiment of the present invention.

4 is a flowchart illustrating a centrifugal separation method according to a fourth embodiment of the present invention.

<Description of the symbols for the main parts of the drawings>

10: Resin film 20: Film separation bag

101: valve member 100: centrifugal separator

110: upper separation bag 111: upper separation space

112: tissue inlet 113: first treatment solution inlet

114: second process solution inlet 115: solution outlet

120: lower separation bag 121: lower separation space

122: purpose discharge port 130: compartment member

Claims (19)

In the centrifugal separation apparatus for separating the target cells from the tissue extracted from the organism using centrifugation, An upper separation space is formed using a flexible material, and a tissue injection port for injecting the extracted tissue into the upper separation space, a treatment solution injection hole for injecting a treatment solution separately from the tissue injection port, and the upper separation after centrifugation. An upper separation bag comprising a solution outlet for discharging residual material in the space; A lower separating bag which is formed integrally with the upper separating bag using a flexible material, forms a lower separating space connected to the upper separating space, and a target discharge port for discharging the target cells from the lower separating space; And A partition member for selectively opening and closing a passage connecting the upper separation space and the lower separation space; Includes, the width of the lower separation space of the lower separation bag is narrower than the width of the upper separation space of the upper separation bag, Separating device for a centrifugal separator characterized in that the width of the upper separation space is gradually reduced corresponding to the width of the lower separation space from a specific point. The method of claim 1, The flexible material is a separator for centrifugation, characterized in that it comprises polyvinyl chloride (polyvinyl chloride). The method of claim 1, The flexible material is a centrifugal separation device using a polymer of dimethylsiloxane, and polycarbonate to selectively permeate gas. The method of claim 1, The upper separation bag includes a valve member for selectively opening and closing the tissue inlet, the treatment solution inlet, and the solution outlet, The lower separating bag comprises a valve member for selectively opening and closing the target outlet port. delete delete The method of claim 1, The partition member is a separator for centrifugation, characterized in that it comprises any one of a forceps, a valve, and a zipper. The method of claim 1, Separating device for centrifugation, characterized in that any one of the tissue inlet, the treatment solution inlet, the solution outlet, and the desired outlet is connected to the outside using a cannula. In the centrifugal separator for separating the target cells from the tissue extracted from the organism using centrifugation, It comprises a film separation bag formed by fusing at least one resin film formed using a synthetic resin, The inside of the film separation bag provides an upper separation space and a lower separation space, the upper portion of the upper separation space tissue injection port for injecting the tissue, treatment solution injection hole for injecting treatment solution separately from the tissue injection port, and centrifugal After the separation is provided with a solution outlet for discharging the remaining material in the upper separation space, the lower portion of the lower separation space is formed with a target outlet for discharging the target cells collected in the lower separation space, A partition member is provided between the upper separation space and the lower separation space to selectively open and close a passage connecting the upper separation space and the lower separation space. The edge of the film separation bag except the tissue inlet, the solution outlet, and the target outlet is sealed by fusion, The width of the upper separation space toward the lower separation space is fused and sealed to gradually decrease from a specific point, Separating device for a centrifugal separation, characterized in that the width of the lower separation space is narrower than the width of the upper separation space. delete 10. The method of claim 9, The edge of the film separation bag is sealed by fusion, and any one of the tissue inlet, the treatment solution inlet, the solution outlet, and the target outlet is connected to the outside using a cannula. Separation device for separation. 10. The method of claim 9, And a valve member for selectively opening / closing the tissue inlet, the treatment solution inlet, the solution outlet, and the target outlet. 10. The method of claim 9, Separating line is formed in the film separation bag along the outer separation space of the lower separation space, the lower separation space is a centrifugal separator for cutting off from the film separation bag in a closed state. The method of claim 13, The boundary between the upper separation space and the lower separation space is separated from the film separation bag in a blocked state by using the partition member. 10. The method of claim 9, The synthetic resin is a centrifugal separation device, characterized in that using a polymer of dimethylsiloxane, and polycarbonate to selectively permeate gas. Claims 1 to 4, 7 to 9, 11 to 15 of any one of the centrifugal separator for separating at least two, One of the centrifugation separator assembly is connected to the tissue inlet of the other centrifugation separator device to increase the capacity of the separation device for principle separation. In the centrifugation method for separating the target cells from the tissue extracted from the organism using centrifugation, An upper separation space is formed using a flexible material, and a tissue inlet for injecting the tissue into the upper separation space, a treatment solution inlet for injecting treatment solution separately from the tissue inlet, and, after centrifugation, in the upper separation space. An upper separation bag including a solution discharge port for discharging residual material, and formed integrally with the upper separation bag by using a flexible material, and forms a lower separation space connected to the upper separation space, and the object from the lower separation space. And a partition member for selectively opening and closing a passage connecting the upper separation space and the lower separation space, the width of the lower separation bag of the lower separation bag to which the discharge outlet is formed, and the upper separation space and the lower separation space. It is formed narrower than the width of the upper separation space of the upper separation bag, Providing a centrifugal separator for gradually dividing the width of the upper separation space corresponding to the width of the lower separation space from a specific point; Injecting the extracted tissue into the upper separation space through the tissue injection port; Injecting a treatment solution reacting with the tissue through the treatment solution inlet into the upper separation space; Placing and centrifuging the upper separating bag and the lower separating bag in the order of centrifugal rotation; Separating the upper separation space and the lower separation space by using the partition member; And Discharging the residual material through the solution outlet; Centrifugal separation method comprising a. The method of claim 17, After discharging the residual material to the solution outlet, And reprocessing the material isolated in the lower separation space using a treatment solution different from the treatment solution. The method of claim 18, The reprocessing step, Connecting a passage connecting the upper separation space and the lower separation space by using the partition member; Injecting the other treatment solution through the treatment solution inlet; Placing and centrifuging the upper separating bag and the lower separating bag in the order of centrifugal rotation; Closing a passage connecting the upper separation space and the lower separation space by using the partition member; And Discharging the residual material through the solution outlet; Centrifugal separation method comprising a.
KR1020090015016A 2009-02-23 2009-02-23 Device for centrifugal separation and method using the same KR101091052B1 (en)

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Cited By (4)

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Publication number Priority date Publication date Assignee Title
KR20200084644A (en) 2019-01-03 2020-07-13 김형섭 Cell separator and method for cell separating using thereof
KR20200084645A (en) 2019-01-03 2020-07-13 김형섭 Cell separator and method for cell separating using thereof
WO2021212192A1 (en) * 2020-04-23 2021-10-28 Inacio Ferreira Junior Celso Bag for collecting and separating blood components
WO2023178498A1 (en) * 2022-03-22 2023-09-28 通用干细胞股份有限公司 Closed cell separation operation apparatus

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KR101313048B1 (en) * 2013-04-09 2013-09-30 김준우 Dividing device for cell fractions or blood elements
KR101343577B1 (en) * 2013-05-03 2013-12-20 김준우 Dividing device for cell fractions or blood elements

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JP2000070763A (en) 1998-09-01 2000-03-07 Toshimasa Yamamoto Separation member, separation vessel and centrifugal separation
JP2002291874A (en) 2001-03-29 2002-10-08 Jms Co Ltd Blood bag for separation of blood components and blood separation method
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JP2000070763A (en) 1998-09-01 2000-03-07 Toshimasa Yamamoto Separation member, separation vessel and centrifugal separation
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20200084644A (en) 2019-01-03 2020-07-13 김형섭 Cell separator and method for cell separating using thereof
KR20200084645A (en) 2019-01-03 2020-07-13 김형섭 Cell separator and method for cell separating using thereof
WO2021212192A1 (en) * 2020-04-23 2021-10-28 Inacio Ferreira Junior Celso Bag for collecting and separating blood components
WO2023178498A1 (en) * 2022-03-22 2023-09-28 通用干细胞股份有限公司 Closed cell separation operation apparatus

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