[go: up one dir, main page]

WO2014201694A1 - Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes - Google Patents

Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes Download PDF

Info

Publication number
WO2014201694A1
WO2014201694A1 PCT/CN2013/077674 CN2013077674W WO2014201694A1 WO 2014201694 A1 WO2014201694 A1 WO 2014201694A1 CN 2013077674 W CN2013077674 W CN 2013077674W WO 2014201694 A1 WO2014201694 A1 WO 2014201694A1
Authority
WO
WIPO (PCT)
Prior art keywords
carrier
frozen
frozen target
target cells
ice
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/CN2013/077674
Other languages
English (en)
Chinese (zh)
Inventor
徐小杨
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.)
Individual
Original Assignee
Individual
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
Application filed by Individual filed Critical Individual
Priority to CN201380003804.0A priority Critical patent/CN104378979B/zh
Priority to PCT/CN2013/077674 priority patent/WO2014201694A1/fr
Publication of WO2014201694A1 publication Critical patent/WO2014201694A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N1/00—Preservation of bodies of humans or animals, or parts thereof
    • A01N1/10—Preservation of living parts
    • A01N1/12—Chemical aspects of preservation
    • A01N1/128—Chemically defined matrices for immobilising, holding or storing living parts, e.g. alginate gels; Chemically altering living parts, e.g. by cross-linking
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N1/00—Preservation of bodies of humans or animals, or parts thereof
    • A01N1/10—Preservation of living parts
    • A01N1/14—Mechanical aspects of preservation; Apparatus or containers therefor
    • A01N1/142—Apparatus
    • A01N1/144—Apparatus for temperature control, e.g. refrigerators or freeze-drying apparatus

Definitions

  • the invention relates to the field of vitrification and freezing technology, in particular to an automatic operation method and system for rapid vitrification of living cells, and a rapid vitrification carrier for living cells. Background technique
  • Rapid vitrification of living cells minimizes interference and damage to cellular biological properties.
  • the frozen fertilized egg or the embryo of the early division can be mechanically cooled, slowly cooled by a programmed cooling device, or fast vitrified freezing.
  • the programmed cooling technique is complicated, the equipment cost is high, the processing time is long, the amount of specimens processed is small, the vitrification is insufficient, and the damage to the cell microstructure is large, and the advantage is that the artificial dependence is small, and a small amount can be achieved.
  • Parallel processing of specimens; rapid vitrification, rapid cooling rate, full vitrification, high recovery rate, but only manual operation, high skill requirements for operators, strict personnel skill training and high operational responsibility, easy Operational accidents have resulted in irregular embryo freezing and even embryo loss.
  • Step 1 Cell dehydration, cryoprotectant balance treatment
  • Step 2 Prepare the pipette tool, cauterize the glass tube, manually stretch, prepare the cell pipette, the inner diameter is less than 1 mm
  • Step 3 Prepare a liquid nitrogen container, fill in a sufficient amount of liquid nitrogen, adjust the microscope at the same time, and prepare the frozen carrier for use
  • Step 4 Manually aspirate the liquid containing the embryonic cell mass from the cryoprotectant using a cell pipette with a manual elastic tip.
  • the amount of liquid required is as small as possible, about 10 to 30 microliters, to stably maintain the position of the liquid column in the cell pipette, the other hand stably holds the frozen carrier, and the liquid containing the embryonic cells is coated on the sheet carrier under a dissecting microscope.
  • the target cells are identified and the excess liquid is aspirated, and the target cells are adhered to the carrier, and the carrier is quickly and manually inserted into the liquid nitrogen to complete the freezing and tube sealing.
  • the main point of this method of operation is, first, the operating time is as short as possible. Living embryos are exposed to the external environment and have a direct effect on cell function. Second, try to smoke as little as possible. Take the liquid and absorb the excess liquid adsorbed on the carrier before the liquid nitrogen is introduced.
  • the purpose is to make the liquid film around the cell as thin as possible, which is beneficial to increase the surface area directly exposed by the cells, thereby increasing the cooling rate.
  • the cooling rate is much higher than the cooling rate required for water crystallization in the cryoprotectant, and vitrification can be achieved.
  • This method of operation relies on the skill of the operator, inefficiency, and the time during which the living embryo is directly exposed to the environment is related to the skill and proficiency of the operator.
  • the operator has a high operational pressure when sucking excess liquid on the carrier.
  • the stability requirements are high and the quality of the refrigeration is unpredictable.
  • This operating system can only process several microliters of specimens at the same time, can not handle a large number of specimens, and severely limits the application range of vitrification technology.
  • Full and rapid completion of the vitrification process is a critical step in freezing living cells and retaining the physiological characteristics and functions of the cells.
  • the existing vitrification technology has a small application range because of the limitation of the freezing carrier, resulting in low freezing efficiency, and only manual processing of small-capacity specimens can not be performed, and it cannot be used for simultaneous processing of large-capacity specimens.
  • the commonly used vitrification technology requires osmotic dehydration treatment of cells, and is equilibrated with a cryoprotectant, and then enters rapid vitrification, but some cells are sensitive to the infiltration process, and osmotic dehydration directly destroys its functional structure, a more reasonable way. It is a direct freezing without cryoprotectant, such as direct vitrification of sperm cell suspension for rapid freezing.
  • Vitrification technology can provide services in important areas such as long-term preservation of important stem cells, resuscitation or donation, and individualized blood component cells.
  • the long-term retention of information on individual cell lines also provides an inexpensive and efficient physical reference frame for cell science and medical physiology research. Therefore, there is a need to develop an automated method and system for rapid vitrification of living cells.
  • Embodiments of the present invention provide an automatic operation method and system for rapid vitrification of living cells, which can quickly and efficiently automate the rapid vitrification of living cells and improve the freezing efficiency.
  • the embodiment of the present invention adopts the following technical solutions:
  • An automatic operation method for rapid vitrification of living cells comprising the steps of:
  • an embodiment of the present invention further provides an automatic operating system for rapid vitrification of living cells, which is capable of performing the above operation method, including a central control device, a mechanical action device, and a freezing medium device and a carrier, wherein the carrier An ice carrier formed by coagulation/deposition of water, and the carrier maintains a specific structure under an operating environment; the central control device acquires a frozen target by reading a chip disposed outside a liquid container containing the frozen target cells The liquid information of the cell is determined according to the liquid information of the frozen target cell, and the carrier corresponding to the liquid information of the frozen target cell is determined, and a corresponding first control command is generated and sent to the mechanical action device; the mechanical action device is Receiving a control command to acquire a corresponding carrier, and taking a liquid containing the frozen target cells from the liquid
  • an embodiment of the present invention further provides a living cell rapid vitrification freezing carrier,
  • the carrier is an ice carrier formed by coagulation/decondensation of water, and the carrier maintains a sheet-like structure under an operating environment, and the carrier is used to carry the surface of the frozen target cells into a specific spatial microstructure.
  • An automatic operation method and system for rapid vitrification of living cells provided by an embodiment of the present invention, by using an ice carrier formed by water solidification/decondensation as a freezing carrier, can quickly adsorb due to the super-hydrophilic property of the ice carrier
  • the liquid rapidly separates the frozen target cells from the liquid, thereby reducing the exposure time of the frozen target cells to the atmosphere, increasing the cooling rate, and reducing the freezing damage.
  • due to the super-adsorption of the ice carrier no need to manually absorb the excess liquid adsorbed on the carrier, so that the automatic operation of the rapid vitrification of the living cells can be realized, the working efficiency can be greatly improved, and the stability of the operation can be ensured. Timeliness and safety. BRIEF DESCRIPTION OF THE DRAWINGS
  • the accompanying drawings which are incorporated in the claims Other drawings may also be obtained from these drawings without the use of creative labor.
  • Embodiment 1 is a flow chart showing an automatic operation method of rapid vitrification of living cells in Embodiment 1 of the present invention
  • Embodiment 2 is a flow chart showing an automatic operation method of rapid vitrification of living cells in Embodiment 2 of the present invention
  • Embodiment 3 is a structural block diagram of an automatic operating system for rapid vitrification of living cells in Embodiment 3 of the present invention
  • Figure 4 is a schematic view showing the structure of a living cell fast vitrification and freezing carrier in the fourth embodiment of the present invention
  • Figure 5 is a schematic view showing the bearing surface structure of a living cell rapid vitrification freezing carrier shown in Figure 4;
  • Fig. 6 is a schematic view showing the structure of a living cell fast vitrification and freezing carrier in Example 5 of the present invention.
  • Figure 7 is a schematic view showing the surface microstructure of the vitrified frozen carrier shown in Figure 6, which can be adjusted by micro The size and spatial shape of the column gaps are used to achieve different microvolume and surface characteristics. detailed description
  • an embodiment of the present invention provides an automatic operation method for rapid vitrification of living cells.
  • the method includes: Step S101: Acquire and freeze the target according to type information of frozen target cells. a carrier corresponding to the type information of the cell; wherein the carrier is an ice carrier formed by water coagulation/desublimation, and the carrier maintains a specific structure under an operating environment; and step S102, placing a liquid containing the frozen target cell On the ice carrier, the liquid is rapidly adsorbed by the ice carrier; step S103, until the liquid film thickness (capacity) of the frozen target cell on the surface of the ice carrier reaches a set thickness (capacity), The ice carrier carrying the frozen target cells is transferred into a freezing medium to complete rapid vitrification.
  • the present invention replaces the human work industry with the automatic operation method of the rapid vitrification of the living cells shown in Fig. 1, which can greatly improve the work efficiency and ensure the stability, timeliness and safety of the operation.
  • Step S201 obtaining a type information of the frozen target cell by reading a chip disposed outside a liquid container storing the frozen target cell;
  • the target cells need to be dehydrated before freezing, and the frozen target cells are placed in a container (injected with a cryoprotectant such as glycerin or DMSO) to balance the cryoprotectant with the frozen target. Free water in the cells to increase the freezing efficiency of the target cells and avoid crystal formation in the target cells.
  • a cryoprotectant such as glycerin or DMSO
  • a chip is provided outside the liquid container containing the frozen target cells, and the chip pre-stores type information of the frozen target cells. Therefore, the type information of the frozen target cells can be obtained by reading a chip provided outside the liquid container in which the frozen target cells are stored.
  • Step S202 comparing the acquired type information of the frozen target cells with pre-stored information, thereby determining and acquiring a carrier corresponding to the type information of the frozen target cells; wherein the carrier is solidified/condensed by water An ice carrier, and the carrier maintains a specific structure in an operating environment;
  • the type information of the different frozen target cells and the correspondence between the carriers may be preset, so that when the type information of the frozen target cells is acquired, the frozen target cells may be obtained by searching the corresponding relationship.
  • the corresponding carrier type may be preset, so that when the type information of the frozen target cells is acquired, the frozen target cells may be obtained by searching the corresponding relationship.
  • the carrier is used to carry a frozen target cell for a freezing operation, which is an ice carrier solidified/condensed by purified water, and the carrier maintains a specific structure in an operating environment.
  • the ice carrier solidified/condensed from pure water is super-hydrophilic and can quickly adsorb liquid.
  • the ice carrier is exposed for a short period of time under operating conditions (e.g., -2 degrees Celsius) sufficient to maintain a substantially fixed sheet structure.
  • the ice carrier is used to carry a surface of the frozen target cell with a specific spatial structure (for example, a porous or surface microscopic structure) to increase the surface area of the carrier adsorbed liquid and improve the adsorption performance.
  • the ice carrier includes a carrier portion and a restriction portion projecting from the carrier portion, the restriction portion for limiting a thickness of a liquid containing the frozen target cells placed on the carrier portion.
  • the specific structure of the carrier will be described in detail later with reference to Figs. Step S203, placing a liquid containing frozen target cells on the ice carrier, so that the liquid is rapidly adsorbed by the ice carrier; Specifically, an appropriate amount of liquid (including frozen target cells) is taken out from the container containing the frozen target cell liquid, and then placed on the corresponding carrier.
  • Step S204 when the liquid film thickness (capacity) of the frozen target cell on the surface of the ice carrier reaches a set thickness (capacity), the ice carrier carrying the frozen target cell is moved into the freezing medium to complete rapid vitrification. Freezing; Specifically, in this step, the following two situations may be included:
  • the frozen target cell is a cell or a collection of cells that need to expose the largest surface area
  • the ice carrier carrying the frozen target cell is immediately transferred into a freezing medium (for example, liquid nitrogen). Rapid vitrification; or
  • the frozen target cell is a cell or a collection of cells that is suitable for simultaneous freezing with an environmental liquid
  • the liquid volume of the frozen target cell to be contained is less than or equal to the capacity determined by the restriction portion
  • the frozen target cell is immediately carried.
  • the ice carrier is transferred into a freezing medium (for example, liquid nitrogen) to complete rapid vitrification.
  • Step S205 performing vacuum freeze-drying treatment on the frozen target cells that have been subjected to rapid vitrification to sublimate the ice carrier carrying the frozen target cells into water vapor, thereby realizing the lyophilized powder state of the frozen target cells. save.
  • a subsequent freeze-drying process is required.
  • human assisted reproductive technology the quality of embryo cryopreservation can be improved.
  • After sperm freezing it can be directly lyophilized under low temperature, and sperm cells can be stored for a long time at a lower cost of lyophilized powder.
  • the specimens can be processed in hundreds of milliliters simultaneously. After the treated specimens are sublimed and vacuumed, the blood components can be stably stored for a long time, and the storage cost and the reprocessing cost are greatly reduced.
  • FIG. 2 is a flow chart showing an automatic operation method for rapid vitrification of living cells according to an embodiment of the present invention. It should be understood that the steps of the automatic operation method for rapid vitrification of living cells proposed by the present invention are not limited to FIG. 2 . In the order of execution shown, those skilled in the art can arbitrarily change the automated method steps of rapid vitrification of living cells in accordance with the teachings of the present invention.
  • the automatic operating system 10 includes a central control device 1, a mechanical action device 2, and a freezing medium device 3 and a carrier 4, wherein the carrier 4 is used to carry frozen target cells for performing a freezing operation, which is An ice carrier formed by solidification/condensation of purified water, and the carrier maintains a specific structure in an operating environment.
  • the ice carrier solidified/condensed from pure water is super-hydrophilic and can quickly adsorb liquid.
  • the ice carrier is maintained in a fixed sheet-like structure under an operating environment (preferably 0 degrees Celsius) to prevent the ice carrier from being melted and deformed.
  • the ice carrier is used to carry the surface of the frozen target cell as a porous structure or other suitable spatial structure, such as a surface microscopic structure having a specific capacity, to increase the surface area of the adsorbed liquid of the carrier, and to improve adsorption. performance.
  • the ice carrier includes a carrier portion and a restriction portion projecting from the carrier portion, the restriction portion for limiting a thickness of the vitrified liquid containing the frozen target cells placed on the carrier portion. The specific structure of the carrier will be described in detail later with reference to Figs.
  • the central control device 1 acquires the vitrified liquid information of the frozen target cells by reading a chip disposed outside the vitrified liquid container containing the frozen target cells, according to the frozen target cells.
  • the vitrification liquid information determines a carrier corresponding to the vitrified liquid information of the frozen target cell, and generates a corresponding first control command to be transmitted to the mechanical action device 2.
  • the central control device 1 is provided with a database, and the database stores different freeze target details. Corresponding relationship between the vitrification liquid information of the cell and the carrier; the central control device 1 searches for the correspondence in the data according to the obtained vitrification liquid information of the frozen target cell, thereby judging the cell with the frozen target
  • the vitrified liquid information corresponds to the type of carrier 4.
  • the mechanical action device 2 acquires a corresponding carrier 4 according to the received first control instruction, and takes out a vitrification liquid containing frozen target cells from the vitrification liquid container, and places the same on the ice carrier.
  • the vitrified liquid is rapidly adsorbed by the ice carrier.
  • the mechanical action device 2 obtains a corresponding frozen carrier according to the received first control command, and then takes out an appropriate amount of liquid (including frozen target cells) from the vitrified liquid container containing the frozen target cells. Placed on the corresponding carrier. Since the carrier is a super-hydrophilic ice carrier, when a vitrification liquid containing frozen target cells is placed on the ice carrier, the vitrification liquid is rapidly adsorbed by the ice carrier, thereby achieving Rapid separation of frozen target cells to expose the frozen target cells.
  • the central control device 1 sends a second control command to the mechanical action device 2 until the liquid film thickness (capacity) containing the frozen target cells on the surface of the ice carrier reaches a set thickness (capacity).
  • the mechanical action device 2 moves the ice carrier carrying the frozen target cells into the freezing medium device 3 (for example, a liquid nitrogen device) to complete rapid vitrification.
  • the freezing medium device 3 for example, a liquid nitrogen device
  • the mechanical action device 2 immediately carries the second control instruction according to the received second control instruction. Freezing the target carrier's ice carrier into a freezing medium (eg, liquid nitrogen) to complete rapid vitrification; or
  • the mechanical action of the vitrified liquid to be included in the frozen target cell is less than or equal to the capacity determined by the restriction portion
  • the device 2 immediately moves the ice carrier carrying the frozen target cells into a freezing medium (for example, liquid nitrogen) according to the received second control command to complete rapid vitrification freezing.
  • a freezing medium for example, liquid nitrogen
  • the automatic operating system further comprises a vacuum freeze-drying device 5, after the ice carrier of the frozen target cell is moved into the freezing medium device 3 to complete rapid vitrification,
  • the central control device 1 sends a third control command to the mechanical action device 2, causing the mechanical action device 2 to move the frozen target cells (along with the carrier) into the vacuum freeze-drying device 5 for vacuum freeze-drying
  • the drying treatment is performed to sublimate the ice carrier carrying the frozen target cells into water vapor, thereby achieving lyophilized powder state preservation of the frozen target cells.
  • a vacuum freeze-drying device 5 is required to perform a subsequent freeze-drying process.
  • human assisted reproductive technology the quality of embryo cryopreservation can be improved.
  • After sperm freezing it can be directly lyophilized under low temperature, and sperm cells can be stored for a long time at a lower cost of lyophilized powder.
  • For the preservation of blood components The specimens in hundreds of milliliters can be processed simultaneously. After the treated specimens are sublimed and vacuumed, the blood components can be stably stored for a long time, and the storage cost and the reprocessing cost are effectively reduced.
  • the automatic operating system for rapid vitrification of living cells of the present embodiment can automatically complete the entire freezing operation process of the frozen target cells by reading the information in the chip provided outside the vitrified liquid container containing the frozen target cells, without Manual participation greatly improves work efficiency and ensures operational stability, timeliness and safety.
  • FIG. 4 a schematic diagram of the structure of a living cell rapid vitrification carrier in Embodiment 4 of the present invention is shown.
  • the carrier 4 of the present embodiment can be used in the above-described automatic operation method and system for rapid vitrification of living cells to carry frozen target cells for a fully automatic freezing operation.
  • the carrier is an ice carrier solidified/condensed from purified water, and the carrier maintains a specific structure in an operating environment.
  • the ice carrier solidified/condensed from pure water is super-hydrophilic and can quickly adsorb liquid.
  • the ice carrier is maintained in a fixed sheet-like structure under an operating environment (e.g., -2 degrees Celsius) to prevent the ice carrier from being melted and deformed.
  • the carrier 4 includes a carrying portion 41 and a restricting portion 42 protruding from the carrying portion 41, and the carrying portion 41 is configured to carry a surface of the frozen target cell as a porous structure 411 (refer to FIG. 5), Increasing the surface area of the carrier adsorbed liquid and improving the adsorption performance.
  • the restricting portion 42 is a plurality of convex protrusions on the carrying portion 41 for limiting the glass containing the frozen target cells placed on the carrying portion 41 Liquid thickness.
  • the carrier 4 provided in this embodiment has super hydrophilic surface characteristics.
  • the hydrophilic material is first perforated, and the size of the pore size is determined by the characteristics of the frozen specimen, so that the aqueous solution can be quickly adsorbed and uniformly distributed into a thin film, thereby rapidly and automatically exposing the target cells, and maximizing the exposed surface area of the target cells.
  • the above structure can enhance the adhesion of the liquid on the carrier, increase the flatness and adhesion of the liquid film, and withstand the acceleration of the mechanical system action, ensuring that the specimen (freezing target cells) is not lost. , to ensure the quality of the frozen.
  • the method of using the vector of the present embodiment is as follows: For freezing of a target cell or a collection of cells that need to expose a maximum surface area, the liquid containing the target cell or cell collection is directly transferred to the functional surface of the carrier, after the target cell is sufficiently exposed, Immediately transferred to a frozen working environment (eg, liquid nitrogen environment); for a cell or collection of cells that are suitable for simultaneous freezing with the culture fluid environment liquid, the liquid volume containing the target cell or collection of cells is less than or equal to the liquid film thickness restriction portion 42 When the carrier capacity is automatically determined, the carrier carrying the target specimen (frozen target cells) is immediately transferred to the frozen working environment to complete the freezing.
  • a frozen working environment eg, liquid nitrogen environment
  • the carrier 4 of the present embodiment can be used in the above-described automatic operation method and system for rapid vitrification of living cells to carry frozen target cells for a fully automatic freezing operation.
  • the carrier is made of a hydrophilic polymer film (e.g., a polycarbonate film material) and is a two-layer composite having a specific surface structure with a layer spacing capable of rapidly adsorbing a liquid (Fig. 7).
  • This embodiment differs from the embodiment 4 in that the carrier 4 is composed of a plurality of carriers of the embodiment 4, and therefore, the carrier of the embodiment is suitable for carrying a large cell or a plurality of target cells for freezing.
  • the carrier of the present invention is composed of different types of carriers by its size, the thickness of the liquid film which is restricted by the restriction portion, and the pore size of the bearing surface, and is thus suitable for different frozen target cells.
  • the automatic operation of rapid vitrification of living cells provided by the embodiments of the present invention is provided.
  • the method and system adopt the ice carrier formed by water solidification/desublimation as a freezing carrier, and the super-hydrophilic property of the ice carrier can rapidly adsorb the liquid, so that the frozen target cells and the liquid are quickly separated, thereby reducing the exposure of the frozen target cells. In the atmospheric environment, increase the rate of cooling and reduce freezing damage.

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Dentistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Environmental Sciences (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

L'invention concerne un procédé automatique de fonctionnement pour la congélation à vitrification rapide de cellules vivantes, comprenant les étapes suivantes : (1) en fonction des informations concernant le type de cellule cible à congeler, l'obtention d'un support correspondant au type de cellule cible à congeler, le support étant un support de type glace obtenu à travers la coagulation/sublimation de l'eau, et le support conservant une structure spécifique dans l'environnement de fonctionnement ; (2) la mise en place du liquide contenant la cellule cible à congeler sur le support de type glace pour permettre au liquide d'être rapidement adsorbé par le support de type glace ; et (3) jusqu'à ce que l'épaisseur du liquide contenant la cellule cible à congeler sur le support de type glace atteigne l'épaisseur cible définie, le transfert du support de type glace portant la cible à congeler dans un milieu de congélation afin d'achever la congélation à vitrification rapide. L'invention concerne également un dispositif automatique de fonctionnement pour la congélation à vitrification rapide de cellules vivantes et un support pour la congélation à vitrification rapide de cellules vivantes.
PCT/CN2013/077674 2013-06-21 2013-06-21 Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes Ceased WO2014201694A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
CN201380003804.0A CN104378979B (zh) 2013-06-21 2013-06-21 活体细胞快速玻璃化冷冻的自动操作方法及系统
PCT/CN2013/077674 WO2014201694A1 (fr) 2013-06-21 2013-06-21 Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/CN2013/077674 WO2014201694A1 (fr) 2013-06-21 2013-06-21 Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes

Publications (1)

Publication Number Publication Date
WO2014201694A1 true WO2014201694A1 (fr) 2014-12-24

Family

ID=52103848

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2013/077674 Ceased WO2014201694A1 (fr) 2013-06-21 2013-06-21 Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes

Country Status (2)

Country Link
CN (1) CN104378979B (fr)
WO (1) WO2014201694A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2017000419A1 (fr) * 2015-06-30 2017-01-05 徐小杨 Procédé et support de congélation/décongélation par vitrification rapide de gouttelette de cellule

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11259520B2 (en) * 2016-08-04 2022-03-01 Fanuc Corporation Stem cell manufacturing system, stem cell information management system, cell transport apparatus, and stem cell frozen storage apparatus

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1821393A (zh) * 2006-03-03 2006-08-23 扬州大学 鸡胚原始生殖细胞的玻璃化冷冻方法
WO2009065585A2 (fr) * 2007-11-20 2009-05-28 MAX-PLANCK-Gesellschaft zur Förderung der Wissenschaften e.V. Dispositif et procédé de congélation ultra-rapide
CN101779623A (zh) * 2010-03-11 2010-07-21 中国农业大学 一种卵母细胞/胚胎玻璃化冷冻保存方法及其冷冻载体
CN102648708A (zh) * 2011-02-25 2012-08-29 深圳华大方舟生物技术有限公司 一种用于胚胎或细胞的冷冻液及其用途
CN102669088A (zh) * 2012-05-30 2012-09-19 河南省农业科学院 哺乳动物胚胎和卵母细胞玻璃化冷冻载体
CN102835389A (zh) * 2012-09-11 2012-12-26 苏州大学 玻璃化冷冻自动化操作系统及操作方法

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001049112A2 (fr) * 2000-01-04 2001-07-12 University Of Connecticut Technique de vitrification d'ovocytes
WO2007018947A2 (fr) * 2005-07-20 2007-02-15 The Regents Of The University Of California Procede et dispositif isochores pour reduire la probabilite d'une nucleation de glace lors de la conservation de matiere biologique a des temperatures inferieures a zero degre centrigrade

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1821393A (zh) * 2006-03-03 2006-08-23 扬州大学 鸡胚原始生殖细胞的玻璃化冷冻方法
WO2009065585A2 (fr) * 2007-11-20 2009-05-28 MAX-PLANCK-Gesellschaft zur Förderung der Wissenschaften e.V. Dispositif et procédé de congélation ultra-rapide
CN101779623A (zh) * 2010-03-11 2010-07-21 中国农业大学 一种卵母细胞/胚胎玻璃化冷冻保存方法及其冷冻载体
CN102648708A (zh) * 2011-02-25 2012-08-29 深圳华大方舟生物技术有限公司 一种用于胚胎或细胞的冷冻液及其用途
CN102669088A (zh) * 2012-05-30 2012-09-19 河南省农业科学院 哺乳动物胚胎和卵母细胞玻璃化冷冻载体
CN102835389A (zh) * 2012-09-11 2012-12-26 苏州大学 玻璃化冷冻自动化操作系统及操作方法

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
ABDELHAFEZ, F.F. ET AL.: "Slow Freezing, Vitrification and Ultra-Rapid Freezing of Human Embryos a Systematic Review and Metaanalysis", REPRODUCTIVE BIOMEDICINE ONLINE, 31 December 2010 (2010-12-31), pages 209 - 222 *
DAVID, H. ET AL.: "A Critical Appraisal of Cryopreservation (Slow Cooling Versus Vitrification) of Human Oocytes and Embryos", HUMAN REPRODUCTION UPDATE, vol. 18, no. 5, 25 April 2012 (2012-04-25), pages 536 - 554 *
QIN, YING ET AL.: "Research Progress on Vitrification and Transplantation of Ovarian Tissue", CHINESE JOURNAL OF TISSUE ENGINEERING RESEARCH, vol. 16, no. 18, 29 April 2012 (2012-04-29), pages 3411 - 3416 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2017000419A1 (fr) * 2015-06-30 2017-01-05 徐小杨 Procédé et support de congélation/décongélation par vitrification rapide de gouttelette de cellule

Also Published As

Publication number Publication date
CN104378979B (zh) 2016-05-11
CN104378979A (zh) 2015-02-25

Similar Documents

Publication Publication Date Title
JP4942896B2 (ja) 新規な凍結試料の加温方法
CN105705626B (zh) 细胞或组织的玻璃化冷冻保存用工具
JP5186676B2 (ja) 細胞の凍結保存のための装置及び方法
JP5825571B2 (ja) 生体試料を凍結保存するための凍結保存用具
US20080176326A1 (en) Delivery of high cell mass in a syringe and related methods of cryopreserving cells
EP3162883B1 (fr) Dispositif et procédé pour la cryoconservation par vitrification de cellules ou de tissus
CN106795475B (zh) 细胞或组织的冷冻保存用夹具及冷冻保存方法
JPH0552451B2 (fr)
US11653644B2 (en) System for rapid cooling and warming of cells and other biological material
WO2014201694A1 (fr) Procédé et système automatiques de fonctionnement pour la congélation à vitrification rapide de cellules vivantes
JP4876922B2 (ja) 生物学的試料保存用デバイス
JP6667903B2 (ja) 中空糸凍結保存用具及び細胞凍結保存方法
WO2017000419A1 (fr) Procédé et support de congélation/décongélation par vitrification rapide de gouttelette de cellule
Austin High-pressure freezing and freeze substitution of Arabidopsis for electron microscopy
JP4849725B2 (ja) 細胞付培養基板及びその製造方法
CN105850986A (zh) 一种高效安全的人稀少精子dpv冷冻系统及冷冻方法
CN204499261U (zh) 一种免支架微型冷冻载体
CN210869621U (zh) 一种螺旋式大规模免疫细胞玻璃化冻存管
JP4214746B2 (ja) 細胞培養基板及びその製造方法
JP2014217356A (ja) 生殖細胞を凍結保存するための自動植氷機能を持った容器の製造方法
CN220004073U (zh) 一种可控开放式的微流控玻璃化冷冻芯片
JP3202440U (ja) 凍結保存用治具
JP7232693B2 (ja) 凍結保存用治具
JP4498260B2 (ja) ガラス化用具のシーリング方法
JP2019187244A (ja) 細胞又は組織の凍結保存用治具

Legal Events

Date Code Title Description
WWE Wipo information: entry into national phase

Ref document number: 201380003804.0

Country of ref document: CN

121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 13887382

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 13887382

Country of ref document: EP

Kind code of ref document: A1