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WO2013147515A1 - Kit de séparation centrifuge et procédé de séparation centrifuge l'utilisant - Google Patents

Kit de séparation centrifuge et procédé de séparation centrifuge l'utilisant Download PDF

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Publication number
WO2013147515A1
WO2013147515A1 PCT/KR2013/002558 KR2013002558W WO2013147515A1 WO 2013147515 A1 WO2013147515 A1 WO 2013147515A1 KR 2013002558 W KR2013002558 W KR 2013002558W WO 2013147515 A1 WO2013147515 A1 WO 2013147515A1
Authority
WO
WIPO (PCT)
Prior art keywords
centrifuge
centrifugation
spin column
container
mixed solution
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/KR2013/002558
Other languages
English (en)
Korean (ko)
Inventor
장기영
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
APINTECH Inc
Original Assignee
APINTECH Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from KR20120079785A external-priority patent/KR101495631B1/ko
Application filed by APINTECH Inc filed Critical APINTECH Inc
Publication of WO2013147515A1 publication Critical patent/WO2013147515A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • 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
    • 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
    • B01L2300/06—Auxiliary integrated devices, integrated components
    • B01L2300/0681—Filter
    • 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
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0848—Specific forms of parts of containers
    • B01L2300/0854—Double walls
    • 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
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0864—Configuration of multiple channels and/or chambers in a single devices comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
    • 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
    • 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
    • B01L2400/0412—Moving fluids with specific forces or mechanical means specific forces centrifugal forces using additionally coriolis forces
    • 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
    • B01L2400/06—Valves, specific forms thereof
    • B01L2400/0622—Valves, specific forms thereof distribution valves, valves having multiple inlets and/or outlets, e.g. metering valves, multi-way valves

Definitions

  • the present invention relates to a centrifugation kit and a method for centrifugation using the same, and more particularly, a sample having a biological or chemical substance such as a nucleic acid is mixed with a buffer to easily collect two different kinds of desired substances through centrifugation. It relates to a centrifugal separation kit and a centrifugation method using the same.
  • the centrifuge kit includes a tubular spin column, a filter inserted under the spin column, and a tubular collection container into which the spin column is inserted.
  • the centrifugation kit is used to inject a mixed solution containing a sample and a buffer for centrifugation in a state in which the spin column and the integration vessel are combined into the spin column, and centrifuge the centrifugation kit in which the spin column and the integration vessel are combined. Loaded in the separator, the centrifugal force of the centrifuge leaves the desired material in the mixed solution contained in the spin column in the filter and the remaining material and buffers are discharged out of the spin column and collected in the integrated vessel.
  • Various types of spin columns are known using this principle (US Pat. No. 4,270,921 (1981), US Pat. No. 4,787,971 (1988), US Pat. No. 6,103,195 (2000), US Pat. 4,871,675 (1989), US Pat. No. 8,062,533 (2011)), commercialized.
  • the present invention has been made to solve the above problems, the centrifugation kit to prevent contamination and loss of the sample during the centrifugation operation, and also to facilitate the separation of the sample, improve workability and facilitate automation And to provide a centrifugal separation method using the same.
  • the present invention provides a mixed solution containing portion containing a mixed solution containing a sample and a buffer having a biological or chemical substance such as nucleic acid for centrifugation, and a centrifuged solution centrifuged from the mixed solution.
  • a spin column formed at a discharge part discharged from the discharge part and a filter part provided at a lower end of the mixed solution receiving part to filter the mixed solution;
  • An integrated container in which a centrifugation liquid accommodating part is formed to receive the centrifuged liquid separated from the spin column; It provides a centrifugal separation kit provided in the centrifugation solution receiving portion of the integrated container, and includes a partition wall that separates the centrifugation liquid receiving portion so as to separate and receive the centrifugal liquid discharged from the discharge portion.
  • the filter unit may be made of a membrane or a media filter.
  • the partition wall is characterized in that it is provided perpendicularly from the inner bottom surface of the integrated container.
  • One side of the integrated container further comprises a secondary integrated container for receiving the integrated container, the centrifugal liquid contained in any one of the centrifugal liquid receiving portion separated by the partition wall flows to the auxiliary integrated container An opening is formed in the inner bottom surface.
  • the step of injecting a mixed solution containing a sample and a buffer having a biological or chemical substance such as nucleic acid for centrifugation to the spin column Loading the integrated vessel into the centrifuge such that the plate surface of the partition is coplanar with the axis of rotation of the centrifuge; Rotating the centrifuge in one direction to collect the centrifuged first centrifuged liquid from the spin column with a centrifugation solution receiving portion located in the opposite direction of rotation of the centrifuge; Stopping the rotation of the centrifuge; Injecting a new buffer different from the buffer into a spin column containing the residual sample remaining by the one-dimensional centrifugation; Rotating the centrifuge in the other direction to collect the centrifuged centrifuge from the spin column to the centrifuge receiving portion located in the opposite direction of rotation of the centrifuge It is also achieved by the centrifugation method used.
  • the present invention it is possible to prevent contamination and loss of the sample during the centrifugal separation operation, and also to facilitate the separation of the sample, to improve the workability, and to facilitate the automation of the centrifugal separation operation.
  • FIG. 1 is an exploded perspective view of a centrifugal separation kit according to a first embodiment of the present invention
  • FIG. 2 is a coupling state diagram of FIG.
  • FIG 3 is a schematic diagram of the centrifugal separation of the centrifugal kit of Figure 1 mounted on a centrifuge rotating in one direction,
  • FIG. 4 is an enlarged view illustrating main parts of FIG. 3;
  • FIG. 5 is a schematic diagram of the centrifugal separation of the centrifugation kit of Figure 1 mounted on a centrifuge rotating in the other direction,
  • FIG. 6 is an enlarged view illustrating main parts of FIG. 5;
  • FIG. 7 is an exploded perspective view of a centrifugal separation kit according to a second embodiment of the present invention.
  • Figure 8 is a schematic diagram of the centrifugal separation of the centrifugal kit of Figure 7 mounted on a centrifuge rotating in one direction,
  • FIG. 9 is a schematic view of the centrifugal separation kit of FIG. 7 mounted on a centrifuge rotating in another direction and centrifuged.
  • FIG. 1 and 2 illustrate a centrifugation kit according to a first embodiment of the present invention.
  • the centrifugation kit 10 includes a spin column 11, a collection container 31, and a partition wall 41. do.
  • the spin column 11 has a predetermined length and has a hollow circular tube shape with openings at both sides.
  • a mixed solution containing a sample containing biological or chemical substances such as nucleic acid for centrifugation and a mixed solution containing a buffer (3,3 ', see FIGS. 3 and 5).
  • the accommodation part 13 is formed inside the spin column 11.
  • One opening of the spin column 11 forms an injection part 15 into which the mixed solution 3, 3 ′ is injected into the mixed solution containing part 13.
  • the cap 17 is mounted to the injection part 15, and the cap 17 is connected to the spin column 11 by a connecting string 19.
  • the cap 17 may be optionally interposed.
  • the other opening of the spin column 11 forms a discharge portion 21 through which centrifugal liquids 5a and 5b are centrifuged from the mixed solution 3 and 3 '.
  • the discharge part 21 forms a step with the mixed solution accommodating part 13 and has a diameter relatively smaller than that of the injection part 15.
  • the filter part 45 is inserted into the lower end of the receiving part 13 of the spin rim 11.
  • the filter part 45 selectively filters the mixed solution 3, 3 'discharged through the discharge part 21, that is, filters out a desired substance from the mixed solution 3,3' containing the sample and the buffer. , Or pass.
  • the filter unit 45 may be formed of a membrane or a media filter. Common membrane components include silica, cellulose, nylon, and PTFE (polytetrafluorethylene). Exchange-exchange resins.
  • the locking column 25 protrudes along the outer circumference of the spin column 11.
  • the catching jaw 25 is an insertion portion of the integrated container 31 which will be described later so that the discharge column 21 of the spin column 11 does not interfere with the partition 41 when the spin column 11 is coupled to the integrated container 31. It is held in 35.
  • the integrated container 31 has a predetermined length and has a hollow circular tube shape with one side opened.
  • a centrifugation liquid receiving portion 33 is formed inside the aggregating vessel 31 to accommodate centrifugal liquids 5a and 5b centrifuged from the spin column 11, and the centrifugal liquid receiving portion 33
  • the spin column 11 is partially insert mounted.
  • the lower region of the centrifugal liquid container 33 is divided into a first centrifugal liquid container 33a and a second centrifugal fluid container 33b by the partition 41.
  • the centrifugal liquid receiving portion located on the left side of the partition 41 of the integrated container 31 is referred to as a first centrifugal liquid receiving portion 33a, as shown in FIG.
  • the centrifugal liquid container is referred to as a second centrifugal fluid container 33b.
  • One opening of the integration container 31 forms an insertion portion 35 into which the spin column 11 is inserted, and the insertion portion 35 communicates with the centrifuge liquid receiving portion 33.
  • a plurality of scales 37 are formed on the outer circumference of the accumulation container 31 so that the amount of centrifugal liquids 5a and 5b contained in the accumulation container 31 can be easily visually checked.
  • the partition 41 has a plate shape and is provided in the lower region of the centrifugal liquid container 33 of the integrated container 31.
  • the partition 41 has a first centrifugal liquid receiving portion 33a and a second centrifugal liquid containing portion at a lower region of the centrifugal liquid receiving portion 33 so as to separate and receive the centrifugal liquid discharged from the discharge portion 21. Separate with (33b).
  • the partition wall 41 has a predetermined height so as not to interfere with the discharge portion 21 of the spin column 11 inserted into the integrated container 31 and has an insertion portion 35 from an inner bottom surface of the integrated container 31. It is erected vertically relative to).
  • a mixed solution 3 containing a sample having a biological or chemical substance such as nucleic acid for centrifugation and a buffer is injected into the spin column 11, and the spin column 11 is inserted into the integration container 31. do.
  • the spin column 11 is inserted into the integrated container 31, and then the mixed solution 3 containing the sample and the buffer having a biological or chemical substance such as nucleic acid for centrifugation and the buffer column is spin column ( 11) may be injected.
  • 3 to 5 show, as an example, a state in which three centrifuge kits 10 according to the present invention are loaded in the centrifuge 1.
  • the centrifuge 1 is rotated in one direction and rotated counterclockwise as shown in FIGS. 3 and 4 for convenience of description.
  • the mixed solution 3 contained in the spin column 11 is centrifuged while being filtered through the filter unit 45 by centrifugal force, and then the centrifuged first centrifuge 5a is spin column 11. Is discharged toward the integrated container 31 through the discharge portion 21 of the.
  • the first centrifuge 5a discharged through the outlet 21 of the spin column 11 flows in the opposite direction of rotation of the centrifuge 1 by inertial force, that is, the clock of the centrifuge 1 Direction is collected by the first centrifugal liquid receiving portion (33a) located in the opposite direction of rotation of the centrifuge (1).
  • the centrifuge 1 is rotated in the other direction, and for the sake of explanation, it is rotated clockwise as shown in FIGS. 5 and 6.
  • the mixed solution (3 ') of the residual sample and the new buffer contained in the spin column 11 is centrifuged while filtered through the filter unit 45 by centrifugal force, and then the centrifuged second centrifuge ( 5b) is discharged toward the integrated container 31 through the discharge portion 21 of the spin column 11.
  • the second centrifugal liquid 5b discharged through the outlet 21 of the spin column 11 flows in the opposite direction of rotation of the centrifuge 1 by inertial force, that is, half of the centrifuge 1. While flowing in the clockwise direction, the second centrifugal liquid receiving portion 33b located in the opposite direction of rotation of the centrifuge 1 is collected.
  • centrifugation kit 10 using the centrifugation kit 10 according to the present invention, two different kinds of desired substances can be easily obtained from a sample having a biological or chemical substance such as nucleic acid.
  • FIG. 7 is shown a centrifugal kit according to a second embodiment of the present invention
  • Figure 8 is a schematic diagram of the centrifugal separation of the centrifugal kit of Figure 7 mounted on a centrifuge rotating in one direction
  • 9 is a schematic view of the centrifugal separation of the centrifugal kit of FIG. 7 mounted on a centrifuge rotating in the other direction.
  • the centrifugal kit 10 ' according to the second embodiment of the present invention further includes an auxiliary integration container 51 for accommodating the integration container 31'.
  • the secondary accumulation container 51 has a predetermined length and has a hollow circular tube shape with one side opened.
  • an auxiliary centrifuge accommodating part 53 for accommodating the one side accommodating part of the integrated container 31 ', that is, the centrifuge 5b centrifuged by the second accommodating part 33b. Is formed, and the integrated centrifuge container 53 is partially inserted into the integrated container 31 '.
  • One opening of the secondary accumulation container 51 forms an accumulation container insertion portion 55 into which the accumulation container 31 'is inserted, and the integration container insertion portion 55 communicates with the auxiliary centrifuge liquid receiving portion 53. .
  • an opening 33c is formed in the inner bottom surface of the integrated container 31' forming the second accommodating portion 33b.
  • the centrifuged liquid 5b centrifuged by the second accommodating portion 33b flows through the opening 33c to the auxiliary centrifugal liquid receiving portion 53 of the auxiliary integrated container 51 as shown in FIG. 9. do.
  • the centrifuge 5a centrifuged by the first accommodating part 33a is collected in the first accommodating part 33a as shown in FIG. 8.
  • the centrifugal separation kit 10 ' according to the second embodiment of the present invention has a large number of centrifugal liquids 5b collected in one receiving portion 33b of the integrated container 31' in which the opening 33c is formed.
  • the buffer can be easily collected in the secondary accumulation container 51.
  • the centrifuge kit according to the present invention by mounting the centrifuge kit according to the present invention to a centrifuge to change the buffer mixed with the sample contained in the kit, and by changing the rotation direction of the centrifuge, it is possible to simply obtain two kinds of the desired material.
  • centrifugal separation kits for example, 96 or 384, may be used in a lattice configuration.
  • Kit 11 Spin Column

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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)
PCT/KR2013/002558 2012-03-30 2013-03-27 Kit de séparation centrifuge et procédé de séparation centrifuge l'utilisant Ceased WO2013147515A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
KR10-2012-0033382 2012-03-30
KR20120033382 2012-03-30
KR20120079785A KR101495631B1 (ko) 2012-03-30 2012-07-23 원심분리 키트 및 이를 이용한 원심분리 방법
KR10-2012-0079785 2012-07-23

Publications (1)

Publication Number Publication Date
WO2013147515A1 true WO2013147515A1 (fr) 2013-10-03

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PCT/KR2013/002558 Ceased WO2013147515A1 (fr) 2012-03-30 2013-03-27 Kit de séparation centrifuge et procédé de séparation centrifuge l'utilisant

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105536900A (zh) * 2016-03-05 2016-05-04 张洪美 一种离心管
WO2016065957A1 (fr) * 2014-10-27 2016-05-06 刘玥萌 Tube de filtre centrifuge
CN105642381A (zh) * 2016-03-05 2016-06-08 张洪美 一种离心管
EP3750994A4 (fr) * 2019-04-19 2021-11-10 Anhui Senpeng Biotechnology Co., Ltd. Procédé de purification et de concentration d'adn dans des échantillons médico-légaux à l'aide d'une colonne de filtration sélective

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6103195A (en) * 1997-08-08 2000-08-15 Shukla; Ashok K. Micro-volume spin columns for sample preparation
US6221655B1 (en) * 1998-08-01 2001-04-24 Cytosignal Spin filter assembly for isolation and analysis
US20020102563A1 (en) * 1998-04-24 2002-08-01 Gjerde Douglas T. Apparatus and method for separating and purifying polynucleotides
US20080020446A1 (en) * 2006-07-22 2008-01-24 Xiyu Jia Plasmid DNA isolation
US8062533B2 (en) * 2008-08-01 2011-11-22 Bioventures, Inc. Devices and methods for the purification, isolation, desalting or buffer/solvent exchange of substances

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6103195A (en) * 1997-08-08 2000-08-15 Shukla; Ashok K. Micro-volume spin columns for sample preparation
US20020102563A1 (en) * 1998-04-24 2002-08-01 Gjerde Douglas T. Apparatus and method for separating and purifying polynucleotides
US6221655B1 (en) * 1998-08-01 2001-04-24 Cytosignal Spin filter assembly for isolation and analysis
US20080020446A1 (en) * 2006-07-22 2008-01-24 Xiyu Jia Plasmid DNA isolation
US8062533B2 (en) * 2008-08-01 2011-11-22 Bioventures, Inc. Devices and methods for the purification, isolation, desalting or buffer/solvent exchange of substances

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2016065957A1 (fr) * 2014-10-27 2016-05-06 刘玥萌 Tube de filtre centrifuge
CN105536900A (zh) * 2016-03-05 2016-05-04 张洪美 一种离心管
CN105642381A (zh) * 2016-03-05 2016-06-08 张洪美 一种离心管
EP3750994A4 (fr) * 2019-04-19 2021-11-10 Anhui Senpeng Biotechnology Co., Ltd. Procédé de purification et de concentration d'adn dans des échantillons médico-légaux à l'aide d'une colonne de filtration sélective

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