WO2016140379A1 - Polymer microcapillary tube for rapid freezing of cells, and microcapillary tube capable of trapping cells - Google Patents

Abstract

According to a polymer microcapillary tube for the rapid freezing of cells, presented in the present invention, by forming a polymer hollow tube, which is made of a polymer material having excellent thermal conductivity, as a polymer microcapillary tube, cells having undergone a cell pretreatment step among the treatment steps in a medical use rapid freezing system for cells can be injected and used in rapid freezing and storage. Meanwhile, according to a microcapillary tube capable of trapping cells, presented in the present invention, by forming a built-in microfilter capable of trapping cells in the microcapillary tube, which is made of a polymer material having excellent thermal conductivity, and enabling the flowing of a cryoprotectant fluid, complex steps of a vitrification process in a medical use rapid freezing system for cells can be simplified and the time required according thereto can be reduced.

Description

Polymer microcapillary for cell freezing and microcapillary with cell trap

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a polymer microcapillary tube for rapid freezing of cells, and more particularly, in a medical cell rapid freezing system consisting of cell pretreatment, cell injection, and rapid freezing and storage, A polymer microcapillary for rapid cell freezing that can be used for rapid freezing and storage after injection.

The present invention also relates to a microcapillary tube capable of trapping cells, and more particularly, to a cell injected for the vitrification process of rapid freezing in a medical cell rapid freezing system consisting of cell pretreatment, cell injection, and quick freezing and storage. A microcapillary tube capable of trapping and allowing cell traps to be used for rapid cooling with fluid control of cryopreservatives.

In general, cell cryopreservation techniques can be broadly divided into two stages: slow freezing and moving rapid cooling (Vitrification, or “Ice Free” cryopreservation). Slow freezing here uses a slower cooling rate than on average 2 ° C./min, resulting in ice formation during the cooling process. These ice crystals act like a blade on cells, causing cell membrane damage and damage to intracellular microstructure, resulting in loss of cell survival as well as normal cell function. In addition, there is a problem in that cell deformation occurs due to a concentration gradient caused by a cryoprotectant during slow cooling.

On the other hand, in the case of rapid freezing, it is possible to implement vitrification (a clear, transparent solid state in which ice does not form, such as glass) while using a low concentration of cryopreservative, thereby reducing cell damage and having a higher survival rate than slow cooling. Here, in order to increase the cooling rate in the rapid freezing, it is necessary to implement a device that maximizes the cooling rate by increasing the temperature difference or decreasing the size of the cooling rate device. Existing rapid cooling devices are not automated by immersing samples containing cells in liquid nitrogen (-196 ° C). Accordingly, there is a need for the development of a one-stop module that integrates the entire process of cryopreservation (cell injection, freezing, storage and thawing).

Accordingly, in order to maximize the cooling rate by reducing the size of the cooling rate device for increasing the cooling rate, a fine capillary device is required. Currently used quartz microcapillary (Quartz microcapillary) has been used in the laboratory level has been published in the paper, but quartz is brittle because of its brittleness is easy to break when cooling, there is a problem that cannot be used in the clinic. That is, Quartz needs additional materials and steps for sealing at both ends in order to go to the closed type.

Prior art 1 (US Pat. No. 6,500,608) is a method invention for vitrification of biological cells, and discloses a method in which cells are directly exposed to a liquid nitrogen. Because of the unknown contaminants in the refrigerant itself and recently living bacteria at very low temperatures, the US FDA does not permit the direct exposure of cells to the refrigerant for cryopreservation of eggs and embryos in artificial fertilization. Prior Art 2 (Vitrification by ultra-fast cooling at a low concentration of cryoprotectants in a quartz micro-capillary: A study using murine embryonic stem cells / Xiaoming He et al / Cryobiology 56 (2008) 223-232) is a quartz capillary (Quartz). capillary) to test the rapid freezing of cells. However, quartz (Quartz) has a good thermal conductivity, but the brittleness is very strong, there is a disadvantage that the cell breaks well during the experiment. In particular, in the case of artificial fertilization (In vitro fertilization) because each cell is important, the broken material has a disadvantage that can never be used in clinical and commercialized.

Accordingly, the present applicant intends to propose a polymer microcapillary tube which is a key component of the medical cell rapid freezing system, which prevents direct exposure of cells, easy sealing at both ends, and prevents cell loss by breaking during cooling. do.

On the other hand, Figure 1 is a view showing the flow of the treatment process in a general medical cell rapid freezing system, Figure 2 is a view showing a conventional vitrification process of rapid cooling. As shown in FIG. 1, a general medical cell rapid freezing system includes a pretreatment of cells, a cell injection process, a quick freezing process, and a storage process. The cryopreserved cells are then thawed later. As shown in Figure 2, the conventional rapid cooling vitrification process, a high concentration (4-8M) cryopreservative is injected into the cell, and the step-by-step injection to recover. That is, the current vitrification process is divided into several stages to inject a high concentration of cryopreservative into the cells, and even in the thawing process after cryopreservation, the vitrification is divided into several stages to remove the cryopreservative in the cells. This stepwise loading and cryopreservation removal process is very complex and time consuming and can lead to fatal damage to cells due to prolonged processing. Meanwhile, many microfluidic devices capable of trapping cells have been developed. However, the microfluidic devices are not suitable for realizing rapid freezing due to their large size and significant drop in thermal conductivity. In addition, microcapillaries that are currently capable of trapping cells have not been implemented.

Prior art 3 (WO2012 / 162779 (International Publication No.)) discloses a microfluidic cell trap and assay device for high throughput analysis, and prior art 4 (Controlled loading of cryoprotectants (CPAs) to oocyte with linear and complex CPA profiles). on a microfluidic platform / H et al / Lab Chip, 2011, 11, 3530 discloses a technique for trapping single oocytes using microfluidic systems and implementing ideal cryopreservant injection trajectories.

The present invention has been proposed to solve the above problems of the conventionally proposed methods, by forming a polymer hollow tube of a polymer material having excellent thermal conductivity as a polymer microcapillary tube, the cell during the treatment in the medical cell rapid freezing system It is an object of the present invention to provide a polymer microcapillary tube for rapid cell freezing, in which cells subjected to a pretreatment procedure can be injected and used for rapid freezing and storage.

In addition, the present invention comprises a polymer hollow tube made of a polymer having excellent thermal conductivity as a polymer microcapillary tube, but having a material and a radius and a thickness for increasing thermal conductivity, so that the cells are not directly exposed, and both ends of the polymer hollow tube It is another object of the present invention to provide a polymer microcapillary tube for quick freezing of cells, which is easy to seal, prevents breakage during the cooling process, prevents cell loss, and allows cells to be vitrified and observed.

In addition, the present invention enables the commercialization of the polymer microcapillary tubes required for rapid cryopreservation techniques, and provides a foundation for the development of integrated automated systems in all processes of cryopreservation. Another object is to provide a polymer microcapillary for quick freezing.

On the other hand, the present invention has been proposed to solve the above problems of the previously proposed methods, trapping the cells in the capillary tube made of a high thermal conductivity polymer material, so that the fluid flow of the cryopreservative It is an object of the present invention to provide a microcapillary tube capable of trapping cells, which can simplify the complicated steps of the vitrification process in the medical cell quick freezing system and reduce the time required.

In another aspect, the present invention provides a microcapillary built-in capillary tube, which allows cell traps, fluid control, and rapid cooling systems to provide an integrated microcapillary tube, while the position of the cell is fixed during handling of the cell during the handling. It is yet another object to provide a cell trap-capable microcapillary tube that does not have to worry about losing cell position, so that it can effectively observe cell volume changes during cryopreservant injection and be used for cell characterization studies.

In addition, the present invention, by having a material, radius and thickness to increase the thermal conductivity of the capillary tube, it is possible to prevent breakage during rapid cooling to prevent cell loss, cell traps and fluid control and microfluidic devices and It is yet another object to provide a cell trap-capable microcapillary, which makes it possible to provide a microcapillary tube that can be integrated as a module of key components of a medical cell rapid freezing system.

Polymer microcapillary for rapid cell freezing according to the characteristics of the present invention for achieving the above object,

Polymer microcapillary for cell deep freezing,

The polymer microcapillary tube,

A polymer hollow tube which forms a hollow in which cells which have been subjected to the cell pretreatment can be injected during the treatment in the medical cell rapid freezing system consisting of cell pretreatment, cell injection, and rapid freezing and storage; And

It includes an opening that is open at both ends of the hollow along the longitudinal direction of the polymer hollow tube (open),

The polymer hollow tube,

After the cell is injected into the hollow, it is characterized by the configuration of the polymer material having a high thermal conductivity so that cracking is prevented even during the treatment of the rapid freezing process.

Preferably, the polymer microcapillary tube,

After the cells are injected into the hollow of the polymer hollow tube, the openings at both ends of the hollow opening may be sealed by heat sealing.

More preferably, the polymer microcapillary tube,

After the openings at both ends of the polymer hollow tube are heat sealed and sealed, the process of rapid freezing and storage may be performed.

Preferably, the polymer microcapillary tube,

The shape of the polymer hollow tube may be configured in a round shape of a cylinder.

More preferably, the polymer hollow tube,

The outer shape may be configured in a round shape of a cylinder, and the hollow of the inner diameter may be configured in a round shape.

Even more preferably, the polymer hollow tube is

The outer shape may be configured in a round shape of a cylinder, but one surface may be configured in a plane to facilitate observation after cell injection.

Even more preferably, the polymer hollow tube is

It is possible to minimize the refraction of light by configuring one surface of the outer shape consisting of a cylindrical round shape in a plane.

Preferably, the polymer microcapillary tube,

The shape of the polymer hollow tube may be configured in a square shape of a square pillar.

More preferably, the polymer hollow tube,

The external shape may be configured in a square shape of a square pillar, and the hollow of the inner diameter may be configured in a square shape.

Even more preferably, the polymer hollow tube is

Through the rectangular shape of the outer surface of the light refraction is minimized to facilitate the observation of the cells injected into the hollow.

Preferably, the polymer microcapillary tube,

For observing the cells injected into the hollow of the polymer hollow tube can be prepared using a photocurable epoxy of a transparent material.

More preferably, the polymer microcapillary tube,

The polymer hollow tube can be produced by an injection molding method.

More preferably, the polymer microcapillary tube,

The polymer hollow tube may be manufactured by a 3D printing process.

More preferably, the polymer microcapillary tube,

The polymer hollow tube may be manufactured by one of injection molding or 3D printing processing methods, but may have a diameter of 500 microns.

Even more preferably, the polymer microcapillary tube,

The polymer hollow tube may have a thickness of 500 microns.

Even more preferably, the polymer microcapillary

The polymer hollow tube may have a diameter of 500 microns and a thickness of 500 microns. However, when the resolution is increased due to improved performance, the polymer hollow tube may be manufactured as a smaller size capillary tube.

Even more preferably, the polymer microcapillary

It is possible to reduce the thickness and the outer diameter of the polymer hollow tube to increase the thermal conductivity of the polymer hollow tube having a thickness of 500micron.

Even more preferably, the polymer microcapillary

Surface mirror processing may be performed by reducing the thickness and outer diameter of the polymer hollow tube to maximize thermal conductivity.

Even more preferably, the polymer microcapillary

Polishing may be performed by reducing the thickness and outer diameter of the polymer hollow tube to maximize thermal conductivity.

Preferably, the polymer microcapillary tube,

The cell is prevented from being directly exposed by injecting the cells into the hollow of the polymer hollow tube, and made of a polymer material having high thermal conductivity, and sealing the openings at both ends by heat sealing, thereby preventing cell loss by breaking during cooling. It can be done.

According to another aspect of the present invention for achieving a further object described above is a microcapillary tube capable of trapping,

A microcapillary tube capable of trapping cells,

Cavities in which cells and cryoprotectants can be injected for vitrification of the rapid freezing process are treated in a medical cell rapid freezing system consisting of cell pretreatment, cell injection, and rapid freezing and storage. A capillary tube having a structure in which hollows at both ends are opened in a longitudinal direction in the form of a hollow tube; And

It characterized in that it comprises a micro filter formed in the hollow of the capillary to trap the cells injected into the hollow formed in the capillary, and to enable the fluid flow of the cryopreservative.

Preferably, the capillary,

It can be made of a polymer material having high thermal conductivity.

Preferably, the capillary,

The cells injected into the hollow may be trapped by the micro filter, and after the cryopreservant is injected, the ends of the hollow may be closed by the sealing portion through heat sealing.

Preferably, the capillary,

In order to observe the cell change injected into the hollow, it may be prepared using a photocurable epoxy of a transparent material.

More preferably, the capillary is

Rapid prototyping (RP) using the photocurable epoxy of the transparent material can be produced.

More preferably, the capillary is

It can be produced by 3D printing processing method.

More preferably, the capillary is

RP (Rapid Prototyping) technology or 3D printing, but one of the manufacturing technology, the diameter of the hollow can be configured to 500㎛.

Even more preferably, the capillary is

The thickness of the capillary tube can be configured to 500㎛.

Even more preferably, the capillary is

It can be configured to a diameter of 500㎛ 500㎛ thickness, but can be composed of a capillary tube of a smaller size (size) when the resolution (resolution) increases due to improved performance.

Preferably, the capillary,

In order to increase the thermal conductivity of the capillary tube having a thickness of 500 μm, the thickness and outer diameter of the capillary tube may be reduced.

More preferably, the capillary is

Surface mirror processing may be performed by reducing the thickness and outer diameter of the capillary tube.

More preferably, the capillary is

Polishing may be performed in a manner that reduces the thickness and outer diameter of the capillary.

More preferably, the capillary is

The hollow tube shape in which the hollow is formed may be configured as one of a round shape of a cylinder or a square shape of a square pillar.

Even more preferably, the capillary is

The outer shape is configured in a round shape of a cylinder, but one surface may be configured in a plane to facilitate observation of the injected cells.

Even more preferably, the capillary is

It is possible to minimize the refractive index of the light by configuring one surface of the outer shape consisting of a cylindrical round shape in a plane.

More preferably, the micro filter,

It may be composed of polystyrene beads (polystyrene beads).

More preferably, the micro filter,

As a structure capable of trapping the cells injected into the hollow of the capillary, the position of the cells is fixed during the cell injection process to maintain the position of the cells during handling, and to observe the volume change of the injected cells during the injection process of the cryopreservative. You can do that.

More preferably, the micro filter,

Styrene bead (Styrene bead) can be stacked in a hollow of the capillary tube and then raised to a specific transition temperature (eg, 100 ℃) to allow each bead to stick to form a filter.

Even more preferably, the micro filter,

The size of the micropores of the filter can be determined by the treatment time and the bead size of the particular transition temperature.

Even more preferably, the micro filter,

The diameter of the polystyrene beads can be configured to 160 μm.

Even more preferably, the micro filter,

Cells injected into the hollow of the capillary are trapped, and the cryopreservant may be configured to form a gap of 20-40 μm with a diameter of the hollow of the capillary so that fluid flow through the polystyrene beads can be controlled. .

According to the polymer microcapillary tube for rapid freezing of cells proposed by the present invention, the polymer microcapillary tube is composed of a polymer hollow tube having excellent thermal conductivity, so that the cell pretreatment process is performed during the treatment in the medical cell rapid freezing system. Cells can be injected and used for quick freezing and storage.

Meanwhile, according to another embodiment of the present invention, the polymer hollow tube made of a polymer having excellent thermal conductivity is composed of a polymer microcapillary tube, and has a material, a radius, and a thickness for increasing thermal conductivity, so that the cells are not directly exposed. It is easy to seal the both ends of the, can be prevented from breaking during the cooling process to prevent cell loss, and to keep the cells in the vitrified state can be observed.

In addition, according to the present invention, it is possible to commercialize the polymer microcapillary tube required for the rapid cryopreservation technology, and to provide a foundation for the development of an integrated automation system in all the processes of cryopreservation. can do.

On the other hand, according to another microcapillary tube trapping proposed in the present invention, by trapping the cells in the capillary tube made of a polymer material having excellent thermal conductivity, by constructing a micro-filter that allows the flow of the cryopreservative fluid in a built-in type In the medical cell rapid freezing system, it is possible to simplify the complicated steps of the vitrification process and to reduce the time required.

In addition, according to the present invention, by constructing a built-in capillary tube of the micro filter, it is possible to provide a micro capillary tube integrated with the cell trap, fluid control, rapid cooling system, the position of the cell is fixed during the cell injection process, the cell during handling There is no risk of losing the location, which allows for effective observation of cell volume changes and use in cell characterization during cryopreservant injection.

In addition, according to the present invention, by having a material, radius and thickness to increase the thermal conductivity of the capillary tube, it is possible to prevent breakage during rapid cooling to prevent cell loss, and to integrate with cell traps and fluid control and microfluidic devices It may be possible to provide a microcapillary tube as a module of key components of a medical cell rapid freezing system.

Figure 1 shows the flow of the process in a typical medical cell rapid freezing system.

2 is a view illustrating a conventional vitrification process of rapid cooling.

Figure 3 is a view showing the flow of processing of the refrigeration system to which the polymer microcapillary tube for rapid cell freezing according to an embodiment of the present invention.

Figure 4 is a view showing the configuration of the processing process of the refrigeration system to which the polymer microcapillary tube for rapid cell freezing according to an embodiment of the present invention.

Figure 5 is a view showing a perspective view of a round shape of the polymer microcapillary for rapid cell freezing according to an embodiment of the present invention.

FIG. 6 is a diagram illustrating a configuration example in which one round surface of a polymer microcapillary tube for rapid freezing of cells is processed in a plane; FIG.

Figure 7 is a view showing the configuration of a rectangular form of the polymer microcapillary for cell rapid freezing according to an embodiment of the present invention.

8 is a view showing a configuration in which the heat sealing of the polymer microcapillary tube for rapid freezing of cells according to an embodiment of the present invention is completed.

Figure 9 is a view showing the configuration of a capillary microcapillary cell trap according to another embodiment of the present invention.

10 is a view showing an example of the configuration processing the outer surface of the round shape in a planar microcapillary possible cell trap according to an embodiment of the present invention.

11 is a view showing an example of the external configuration of the cell trap is possible microcapillary tube according to an embodiment of the present invention.

12 is a view showing a state of use of the microcapillary capable of trapping cells according to another embodiment of the present invention.

<Description of the code>

100: polymer microcapillary tube according to an embodiment of the present invention

101: hollow 102: polymer hollow tube

103: opening 104: heat seal

110: capillary 111: hollow

112: sealing portion 120: micro filter

Hereinafter, exemplary embodiments of the present invention will be described in detail with reference to the accompanying drawings so that those skilled in the art may easily implement the present invention. However, in describing the preferred embodiment of the present invention in detail, if it is determined that the detailed description of the related known function or configuration may unnecessarily obscure the subject matter of the present invention, the detailed description thereof will be omitted. In addition, the same reference numerals are used throughout the drawings for parts having similar functions and functions.

In addition, in the specification, when a part is 'connected' to another part, it is not only 'directly connected' but also 'indirectly connected' with another element in between. Include. In addition, the term "comprising" a certain component means that the component may further include other components, except for the case where there is no contrary description.

3 is a view showing the flow of the process of the refrigeration system to which the polymer microcapillary tube is applied for rapid cell freezing according to an embodiment of the present invention, Figure 4 is a rapid cell freezing according to an embodiment of the present invention Figure showing the configuration of the treatment process of the refrigeration system to which the polymer microcapillary tube is applied. As shown in Figures 3 and 4, the refrigeration system to which the polymer microcapillary tube for rapid freezing of cells according to an embodiment of the present invention is applied, is treated by the process of cell pretreatment, cell injection, rapid freezing, and storage Will be made. Hereinafter, the polymer microcapillary tube 100 that can be used for cell injection, rapid freezing, and storage after the cell pretreatment is performed in the medical cell rapid freezing system will be described in detail.

Figure 5 is a view showing a perspective view of the round shape of the polymer microcapillary for rapid cell freezing according to an embodiment of the present invention, Figure 6 is a polymer microcapillary for rapid cell freezing according to an embodiment of the present invention FIG. 7 is a view showing an example of a configuration in which one surface of a round shape is processed into a plane, and FIG. 7 is a diagram illustrating a rectangular configuration of a polymer microcapillary tube for rapidly freezing cells according to an embodiment of the present invention. As shown in FIGS. 5 to 7, the polymer microcapillary tube 100 for rapid cell freezing according to an embodiment of the present invention may include a polymer hollow tube 102 and an opening part forming a hollow 101. 103, and may further include a heat seal (104).

The polymer hollow tube 102 is a hollow 101 into which cells which have been subjected to cell pretreatment can be injected during the treatment in a medical cell rapid freezing system consisting of cell pretreatment, cell injection, and rapid freezing and storage. ) Is a constitution of the microtubules. The polymer hollow tube 102 is preferably made of a polymer material having high thermal conductivity so that the cell can be prevented from being broken even when the cell is injected into the hollow 101 during the rapid freezing process. Here, as shown in FIG. 5, the polymer hollow tube 102 of the polymer microcapillary tube 100 may have a polymer shape in a cylindrical round shape. At this time, the polymer hollow tube 102 constituting the outer shape in a round shape of a cylinder may be configured to form a hollow 101 of the inner diameter in a round shape. In addition, the polymer hollow tube 102 having an outer shape in a cylindrical round shape may have a flat surface on one side to facilitate observation after cell injection. Here, in the polymer hollow tube 102, the reason for forming a flat surface of a cylindrical shape in a cylindrical shape is to minimize the refraction of light to facilitate cell observation.

Meanwhile, as shown in FIG. 7, the polymer hollow tube 102 may have a polymer shape in a square shape of a square pillar. At this time, the polymer hollow tube 102 constituting the outer shape in a square shape of the square pillar is to configure the hollow 101 of the inner diameter in a square shape. The polymer hollow tube 102 may be made to easily observe the cells injected into the hollow 101 by minimizing the refraction of light through the rectangular shape of one surface.

The opening part 103 is a structure in which the hollows 101 at both ends are opened along the longitudinal direction of the polymer hollow tube 102. That is, the open part 103 refers to a portion in which the hollows 101 are opened at both ends along the length direction of the polymer hollow tube 102. The opening 103 at both ends is sealed through a heat seal 104 which will be described later.

The heat seal 104 is configured to seal both ends of the polymer hollow tube 102 after cells are injected into the hollow 101 of the polymer hollow tube 102. Here, the treatment of heat sealing 104 is a treatment configuration that requires additional materials and steps for sealing at both ends in order to go from the conventional quartz capillary to the closed type. To be resolved.

The polymer microcapillary tube 100 is injected into the hollow 101 of the polymer hollow tube 102 to prevent the cells from being directly exposed, and is composed of a polymer material having high thermal conductivity, and the amount of heat sealing 104. The opening 103 at the tip can be sealed so that it can not be broken during the cooling process so that cell loss can be prevented. As shown in FIG. 8, the polymer microcapillary tube 100 is hollow 101 through a heat sealing process after cells are injected into the hollow 101 of the polymer hollow tube 102. The openings 103 at both ends of the open side can be sealed. Here, the polymer microcapillary tube 100 may be used to perform a process of rapid freezing and storage after the openings 103 at both ends of the polymer hollow tube 102 are heat sealed 104 and sealed.

In addition, the polymer microcapillary tube 100 may be manufactured using a photocurable epoxy of a transparent material for observation of cells injected into the hollow 101 of the polymer hollow tube 102. In this case, the polymer microcapillary tube 100 may be manufactured by the injection molding method of the polymer hollow tube 102, or may be manufactured by the 3D printing processing method of the polymer hollow tube 102. In addition, the polymer microcapillary tube 100 is manufactured by one of the injection or 3D printing processing method of the polymer hollow tube 102, the diameter may be configured to 500micron. In addition, the polymer microcapillary tube 100 may be configured to have a thickness of 500 micron of the polymer hollow tube 102.

In addition, the polymer microcapillary tube 100 is composed of a diameter of 500 micron and a thickness of 500 micron of the polymer hollow tube 102, but when the resolution is increased due to the improvement in performance, the microcapillary tube of smaller size may be manufactured. Can be. Here, the polymer microcapillary tube 100 may reduce the thickness and outer diameter of the polymer hollow tube 102 to increase the thermal conductivity of the polymer hollow tube 102 having a thickness of 500 micron. In this case, the polymer microcapillary tube 100 may perform surface mirror processing in such a manner as to reduce the thickness and outer diameter of the polymer hollow tube 102 to maximize thermal conductivity. In addition, the polymer microcapillary tube 100 may be polished in such a manner as to reduce the thickness and outer diameter of the polymer hollow tube 102 to maximize thermal conductivity.

Figure 5 is a polymer microcapillary tube 100 for rapid freezing of cells according to an embodiment of the present invention, shows a perspective view of a polymer hollow tube 102 configured in a round shape of a cylinder. FIG. 6A illustrates a side view of one surface of a polymer hollow tube 102 having a cylindrical round shape, and FIG. 6B shows a polymer hollow tube 102 having a cylindrical round shape. In Fig. 1, a perspective view in which one surface is configured as a plane is shown. 7 is a polymer microcapillary tube 100 for rapid freezing of cells according to an embodiment of the present invention. 8 shows a configuration example in which the heat seal 104 of the polymer microcapillary tube 100 for rapid freezing of cells according to an embodiment of the present invention is formed.

9 is a view showing the configuration of a capillary microcapillary cell trap according to another embodiment of the present invention. As shown in FIG. 9, the microcapillary tube capable of trapping cells according to another embodiment of the present invention may include a capillary tube 110 and a micro filter 120.

The capillary 110 is a cell and cryoprotectant to be injected for the vitrification process of rapid cooling in the medical cell rapid freezing system consisting of cell pretreatment, cell injection and rapid freezing and storage. It is a configuration to have a structure in which the hollow 111 at both ends in the longitudinal direction in the form of a hollow tube to form a hollow 111 that can be opened. The capillary tube 110 may be made of a polymer material having high thermal conductivity so that the cell can be prevented from being broken even when the cell is injected into the hollow 111 during the rapid freezing process. In addition, the capillary 110 is trapped by the micro-filter 120 which will be described later the cells injected into the hollow 111, the hollow 111 is treated by heat sealing after the cryopreservant is injected. Both ends of the open end may be finished with the seal 112.

In addition, the capillary 110 may be manufactured using a photocurable epoxy of a transparent material for the observation of cell changes injected into the hollow 111. In this case, the capillary tube 110 may be a hollow tube made by Rapid Prototyping (RP) technology as a method of using a photocurable epoxy of a transparent material. The advantage of RP is that we can freely make the shape we want, and by using the photocurable epoxy, the light transmittance of the material is excellent, so that the experimenter can easily observe the cells injected into the capillary 110, and also with the help of other complicated devices It is possible to implement the structure of the micro filter 120 in the fine capillary without receiving. In addition, the capillary 110 may be manufactured by a 3D printing processing method, in addition to the approach of the RP technology. At this time, the capillary 110 is manufactured by one of RP (Rapid Prototyping) technology or 3D printing processing technology, the diameter of the hollow 111, that is, the inner diameter can be configured to 500㎛, the thickness of the capillary 110 500㎛ It can be configured as.

In addition, the capillary 110 may be configured to have a diameter of 500 μm × 500 μm, but may also be configured as a capillary 110 having a smaller size when resolution is increased due to performance improvement. The capillary 110 may be further made to reduce the thickness and outer diameter of the capillary 110 to increase the thermal conductivity of the capillary 110 having a thickness of 500 μm. Here, the capillary tube 110 may be subjected to a surface mirror surface treatment process to reduce the thickness and outer diameter of the capillary tube 110, and another method may be a polishing process to reduce the thickness and outer diameter of the capillary tube 110. May be

Meanwhile, as illustrated in FIG. 11, the capillary tube 110 may be configured to have a hollow tube shape in which the hollow 111 is formed in one of a round shape of a cylinder or a square shape of a square column. Here, the capillary 110 can be implemented in various vessel models through 3D printing or RP technology. In addition, as shown in FIG. 10, the capillary 110 may be configured to have one surface in a plane to facilitate observation of the injected cells when forming the outer shape in a round shape of a cylinder. The capillary 110 may be configured to form a flat surface of a cylindrical shape in a round shape, thereby minimizing the refractive index of the light to facilitate the observation of the injected cells.

The micro filter 120 traps cells injected into the hollow 111 formed in the capillary 110, and is formed in the hollow 111 of the capillary 110 to allow the fluid flow of the cryopreservative. . That is, the micro filter 120 is a structure capable of trapping the cells injected into the hollow 111 of the capillary 110, the position of the cells is fixed during the cell injection process to maintain the position of the cells during handling, cryopreservative During the injection process, the volume change of the injected cells can be observed. The micro filter 120 may be composed of polystyrene beads. Here, the micro filter 120 is formed by stacking styrene beads in the hollow 111 of the capillary 110 and stacking the beads at a specific transition temperature (eg, 100 ° C.) to form respective filters. Do it. In this case, the size of the micro holes of the filter may be determined according to the processing time and the bead size of the specific transition temperature. In addition, the micro filter 120 may preferably be configured to a diameter of 160 ㎛ polystyrene beads. In this case, as shown in FIG. 12, the micro filter 120 traps cells injected into the hollow 111 of the capillary tube 110, and the cryopreservative may control the flow of the fluid through the polystyrene beads. It is possible to form a gap (Gap) of 20-40㎛ the diameter of the hollow 111 of 110.

FIG. 10 is a diagram illustrating a configuration example in which a rounded outer surface of a microcapillary tube according to an embodiment of the present invention is processed in a plane. FIG. 10A illustrates a side view of one surface of the capillary tube 110 having a cylindrical round shape, and FIG. 10B illustrates one surface of the capillary 110 having a cylindrical round shape. The perspective view structure comprised in a plane is shown. That is, the capillary tube 110 as shown in FIG. 10 is configured to form a surface of a cylindrical round shape in a plane to minimize the refractive index of light to facilitate observation of the injected cells.

11 is a view showing an example of the external configuration of a microcapillary tube capable of trapping cells according to an embodiment of the present invention. FIG. 11A illustrates a capillary tube capable of trapping cells, and shows an example in which the external shape of the capillary tube 110 in which the microfilter 120 is formed in the hollow 111 in the form of a cylinder is rounded. (B) shows an example in which the external shape of the capillary tube 110 in which the microfilter 120 is formed in the hollow 111 as a microcapillary tube capable of trapping cells is formed in a square column shape. In addition, the capillary 110 may be implemented in various vessel models through 3D printing or RP technology.

12 is a view showing the state of use of the microcapillary capable of trapping cells according to an embodiment of the present invention. 12A illustrates an example in which a capillary tube 110 and a micro static mixing tee are connected to each other, and FIG. 12B illustrates a cell injected into the capillary tube 110 in a visceral form. The structure of the state where the cell was trapped by the micro filter 120 which becomes is shown. That is, as shown in FIG. 12, the capillary tube 110 in which the micro filter 120 is formed in a built-in shape holds cells (eg, eggs) which are connected to a micro static mixing tee and injected. The cryopreservative may be passed through the polystyrene beads of the micro filter 120 to enable fluid flow.

As described above, the cell trap-capable microcapillary tube according to an embodiment of the present invention solves the problem that the conventional microfluidic system does not allow rapid freezing during cryopreservation after cell treatment. Integration with cryopreservant injection technology enables the development of an integrated refrigeration system with an integrated microfluidic device with automatic cell pretreatment. In addition, the capillary tube 110 of the built-in filter obtained by the RP technology is used for rapid cooling, as well as can be used in the study of cell characteristics by observing the change of the cell according to the cryopreservant injection profile of the cells in the prototype form.

The present invention described above may be variously modified or applied by those skilled in the art, and the scope of the technical idea according to the present invention should be defined by the following claims.

Claims (41)

Polymer microcapillary for cell deep freezing, The polymer microcapillary tube 100, Polymer hollow tube forming a hollow 101 through which cell pretreatment can be introduced during the treatment in a medical cell rapid freezing system consisting of cell pretreatment, cell injection, and rapid freezing and storage. 102); And Including the opening 103 is open at both ends of the hollow 101 in the longitudinal direction of the polymer hollow tube 102, The polymer hollow tube 102, After the cell is injected into the hollow 101, characterized in that the thermal conductivity of the polymer material is configured so that the crack is prevented even during the processing of the rapid freezing process, polymer microcapillary for cell rapid freezing. The method of claim 1, wherein the polymer microcapillary tube 100, After the cells are injected into the hollow 101 of the polymer hollow tube 102, the openings 103 at both ends of which the hollow 101 is opened through the heat sealing 104 are sealed. Characterized in the polymer microcapillary for cell rapid freezing. The method of claim 2, wherein the polymer microcapillary tube 100, After the openings (103) at both ends of the polymer hollow tube (102) is heat sealed (104) is sealed, characterized in that the process of rapid freezing and storage is carried out, polymer microcapillary for cell rapid freezing. The method of claim 1, wherein the polymer microcapillary tube 100, The shape of the polymer hollow tube (102) is characterized in that the cylindrical round shape of the polymer, capillary polymer microcapillary tube for rapid freezing. The method of claim 4, wherein the polymer hollow tube 102, A cylindrical microcapillary tube for rapid freezing of cells, characterized in that the outer shape is formed in a cylindrical shape, and the hollow 101 of the inner diameter is formed in a round shape. The method of claim 5, wherein the polymer hollow tube 102, A cylindrical microcapillary tube for rapid freezing of cells, characterized in that the outer shape is configured in a round shape, but is configured in one plane in order to facilitate observation after cell injection. The method according to claim 6, wherein the polymer hollow tube 102, A polymer microcapillary tube for rapid freezing of cells, characterized by minimizing the refraction of light by forming a flat surface of the outer shape consisting of a cylindrical round shape. The method of claim 1, wherein the polymer microcapillary tube 100, The shape of the polymer hollow tube (102) is characterized in that consisting of a rectangular column of square pillars, polymer microcapillary for rapid cell freezing. The method of claim 8, wherein the polymer hollow tube 102, Comprising an external shape in the form of a square of square pillars, characterized in that the hollow 101 of the inner diameter in the form of a square, polymer microcapillary for rapid cell freezing. The method of claim 9, wherein the polymer hollow tube 102, Minimized refraction of light through the rectangular shape of the outer surface to facilitate observation of the cells injected into the hollow 101, polymer microcapillary for rapid cell freezing. The method according to any one of claims 1 to 10, wherein the polymer microcapillary tube 100, In order to observe the cells injected into the hollow 101 of the polymer hollow tube 102, characterized in that the production using a photocurable epoxy of a transparent material, polymer microcapillary for rapid cell freezing. The method of claim 11, wherein the polymer microcapillary tube 100, The polymer hollow tube (102), characterized in that produced by the injection molding method, polymer microcapillary tube for rapid cell freezing. The method of claim 11, wherein the polymer microcapillary tube 100, The polymer hollow tube (102) is characterized in that produced by the 3D printing (printing) processing method, polymer microcapillary tube for rapid cell freezing. The method of claim 11, wherein the polymer microcapillary tube 100, The polymer hollow tube 102 is manufactured by one of injection or 3D printing processing methods, but characterized in that the diameter of 500micron, polymer microcapillary tube for rapid freezing of cells. The method of claim 14, wherein the polymer microcapillary tube 100, Polymer micro capillary for cell rapid freezing, characterized in that the polymer hollow tube 102 is configured to a thickness of 500micron. The method of claim 15, wherein the polymer microcapillary tube 100, The polymer hollow tube 102 is configured to have a diameter of 500 microns and a thickness of 500 microns, but when the resolution is increased due to improved performance, the microcapillary tube having a smaller size is produced. Polymer microcapillary for application. The method of claim 15, wherein the polymer microcapillary tube 100, A polymer microcapillary tube for rapid cell freezing, characterized in that to reduce the thickness and outer diameter of the polymer hollow tube (102) to increase the thermal conductivity of the polymer hollow tube (102) having a thickness of 500micron. The method of claim 17, wherein the polymer microcapillary tube 100, The surface of the polymer hollow tube (102) by reducing the thickness and outer diameter as possible to increase the thermal conductivity, characterized in that to perform a surface mirror surface treatment, polymer microcapillary for cell rapid freezing. The method of claim 17, wherein the polymer microcapillary tube 100, The polymer microcapillary tube for rapid freezing of cells, characterized in that the polishing (polishing) is carried out in such a way as to reduce the thickness and outer diameter of the polymer hollow tube (102) to maximize the thermal conductivity. The method of claim 1, wherein the polymer microcapillary tube 100, The cells are injected into the hollow 101 of the polymer hollow tube 102 to prevent the cells from being directly exposed, and are formed of a polymer material having a high thermal conductivity, and the open portions 103 at both ends thereof are heat sealed 104. A polymer microcapillary for cell rapid freezing which is sealed so that it does not break during the cooling process to prevent cell loss. A microcapillary tube capable of trapping cells, Cells and cryoprotectants may be injected for vitrification during rapid pretreatment in a medical cell rapid freezing system consisting of cell pretreatment, cell injection, and rapid freezing and storage. A capillary tube 110 having a structure in which hollows 111 at both ends are opened in a longitudinal direction in the form of a hollow tube to form 111; And It includes a micro filter 120 is formed in the hollow 111 of the capillary 110 to trap the cells injected into the hollow 111 formed in the capillary 110, the fluid flow of the cryopreservative. A microcapillary tube capable of trapping cells. The method of claim 21, wherein the capillary 110, A microcapillary tube capable of trapping cells, characterized in that it is composed of a polymer material having high thermal conductivity. The method of claim 21, wherein the capillary 110, Cells injected into the hollow 111 are trapped by the micro filter 120, and after the cryopreservant is injected, both ends of the hollow 111 are opened through a heat sealing process. Microcapillary capable of trapping cells, characterized in that it is finished with 112). The method of claim 21, wherein the capillary 110, Cell trap capable of fine capillary tube, characterized in that the production using a transparent photocurable epoxy for observation of cell changes injected into the hollow (111). The method of claim 24, wherein the capillary 110, Cell trap-capable microcapillary tube, characterized in that produced by Rapid Prototyping (RP) technology using the photocurable epoxy of the transparent material. The method of claim 24, wherein the capillary 110, A microcapillary tube capable of trapping a cell, which is produced by a 3D printing processing method. The method of claim 24, wherein the capillary 110, RP (Rapid Prototyping) technology or 3D printing, but one of the manufacturing technology, characterized in that the hollow 111 is characterized in that the diameter of 500㎛, capillary cell trap capable. The method of claim 27, wherein the capillary 110, The capillary 110 is characterized in that the thickness of 500㎛, capillary cell trap capable. The method of claim 28, wherein the capillary 110, 500 μm in diameter × 500 μm in thickness, but when the resolution is increased due to improved resolution, the microcapillary tube trap is possible. The method according to any one of claims 21 to 29, wherein the capillary 110, To increase the thermal conductivity of the capillary tube (110) having a thickness of 500㎛, characterized in that to reduce the thickness and outer diameter of the capillary tube (110), cell trap capable of capillary. The method of claim 30, wherein the capillary 110, The surface capacitive processing is performed in a way to reduce the thickness and outer diameter of the capillary 110, the cell trap is possible microcapillary. The method of claim 30, wherein the capillary 110, The capillary 110, characterized in that the polishing (polishing) is carried out by reducing the thickness and outer diameter of the capillary, cell trap capable of capillary. The method of claim 30, wherein the capillary 110, The hollow tube shape formed with the hollow 111 is characterized in that it comprises one of a round shape of a cylinder, or a rectangular shape of a square column, a cell trap capable of capillary. The method of claim 33, wherein the capillary 110, Comprising a round shape of a cylindrical shape, characterized in that one surface is configured in a plane to facilitate observation of the injected cells, the cell trap is possible microcapillary tube. The method of claim 34, wherein the capillary 110, Cell trapping is possible microcapillary, characterized in that by minimizing the refractive index of the light by forming a plane of the outer surface consisting of a cylindrical round shape. The method of claim 30, wherein the micro filter 120, A microcapillary tube capable of trapping a cell, characterized in that it is composed of polystyrene beads. The method of claim 30, wherein the micro filter 120, As a structure capable of trapping cells injected into the hollow 111 of the capillary 110, the position of the cells is fixed during the cell injection process to maintain the position of the cells during handling, and the cells injected during the injection of cryopreservative Cell capping is possible microcapillary, characterized in that the volume change can be observed. The method of claim 30, wherein the micro filter 120, Cell trap-capable microcapillary, characterized in that the styrene beads (Styrene bead) is stacked in the hollow of the capillary tube and then raised to a specific transition temperature (for example 100 ℃) to each bead to form a filter. The method of claim 38, wherein the micro filter 120, A microcapillary capable of trapping cells, characterized in that the size of the micropores of the filter is determined by the treatment time and the bead size of a particular transition temperature. The method of claim 36, wherein the micro filter 120, A fine capillary tube capable of trapping cells, characterized in that the diameter of the polystyrene beads is 160 µm. The method of claim 40, wherein the micro filter 120, Cells injected into the hollow 111 of the capillary 110 are trapped, and cryopreservatives are 20-40 μm in diameter with the diameter of the hollow 111 of the capillary 110 so that fluid flow through the polystyrene beads can be controlled. Forming a gap (Gap) of, Capillary capillary cell trap capable.

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