KR20140117944A - Method of fabriciationg a membrane structure and an apparatus for culturing comprising the same - Google Patents

Abstract

A cell culture apparatus according to an embodiment of the present invention includes a lower structure including a culture medium storage portion for storing a culture medium, a culture medium injection port connected to the culture medium storage portion and supplying the culture medium, And a membrane for absorbing the culture solution from the culture medium reservoir through a plurality of pores adjacent to the membrane structure; and a membrane electrode assembly including a cell inlet for providing cells to the membrane on the membrane structure, And an upper structure having a cell outlet.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention [0001] The present invention relates to a method for manufacturing a membrane structure and a cell culture apparatus using the membrane structure,

The present invention relates to a method for producing a microporous membrane structure and a cell culture apparatus using the membrane structure.

Cell culture is generally aseptically removed from a multicellular organism and cultured and propagated in the vessel. According to the cell culture method, the cell culture is divided into single cell culture, small organ culture, plant tissue culture, and animal tissue culture. Cell culture is performed in various fields such as production of antivirus, development of new drugs, development of artificial organs, production of medicines by gene manipulation of animal cells, or breeding by cell fusion of plants.

Cell culture requires a certain space in which cells can be cultured, a culture medium that supplies nutrients to the cells, and a culture storage space that stores the culture medium. In particular, the culture medium, the gaseous substance, and the stimulating substance are injected into the culture medium storage space and used in the cell culture, and then exchanged at appropriate intervals to keep the cell tissue fresh.

The present invention relates to a method for producing a membrane structure capable of supplying an effective culture medium to cells.

Another object of the present invention is to provide a cell culture apparatus including a membrane structure capable of supplying an effective culture liquid.

The problems to be solved by the present invention are not limited to the above-mentioned problems, and other problems not mentioned can be clearly understood by those skilled in the art from the following description.

A cell culture apparatus according to an embodiment of the present invention includes a lower structure including a culture medium storage portion for storing a culture medium, a culture medium injection port connected to the culture medium storage portion and supplying the culture medium, And a membrane for absorbing the culture solution from the culture medium reservoir through a plurality of pores adjacent to the membrane structure; and a membrane electrode assembly including a cell inlet for providing cells to the membrane on the membrane structure, And an upper structure having a cell outlet.

The upper structure may further include a partition disposed on the membrane and partitioning the cell culture section.

The side walls of the membrane exposed to the septum may have a vertical, tapered, or concave shape from the lower surface of the membrane.

The membrane structure may further include a support for supporting the membrane, the support structure being disposed on both sides of the culture solution reservoir and on the lower structure, and an etch stop pattern interposed between the support and the membrane.

The membrane is a 1? To 20 < RTI ID = 0.0 >. ≪ / RTI >

The pores are 0.5? To < RTI ID = 0.0 > 20. ≪ / RTI >

The membrane may comprise a silicon on insulator (SOI), silicon oxide, silicon nitride, a combination of silicon oxide and silicon nitride, a polymeric material, a glass or a metal material.

A method of fabricating a membrane structure according to an embodiment of the present invention includes forming an etch stop layer on a semiconductor substrate, forming a membrane layer on the etch stop layer, patterning the membrane layer to expose an upper surface of the etch stop layer, Etching the lower surface of the semiconductor substrate to form a support for exposing a portion of the etch stop layer; and patterning the etch stop layer exposed on the support to etch through the holes, Preventing pattern.

Removing the etch stop pattern to completely expose the surface of the membrane after forming the etch stop pattern; And hydrophilically surface-modifying the surface of the membrane.

Hydrophilic surface modification may include oxygen plasma treatment of the membrane surface or coating of a chemical agent containing a surfactant.

The etch stop layer may be formed of a material having etch selectivity with the semiconductor substrate.

The etch stop layer may be formed of a material having hydrophilicity.

The membrane membrane is a 1? To 20 < RTI ID = 0.0 >. ≪ / RTI >

The membrane may be formed by performing a wet etching or a dry etching process.

In the cell culture apparatus according to an embodiment of the present invention, the culture liquid storage portion storing the culture liquid is disposed below the cell culture portion provided with the cells. The culture solution is supplied from the culture solution storage part to the cell culture part through an ultra thin membrane, and the cells can be supplied with a uniform culture solution.

In addition, the cell may be allowed to react with the culture liquid at the cell outlet, thereby discharging the generated waste. Therefore, it is possible to prevent contamination of the culture liquid in the cell culture apparatus, and to proliferate the cells under conditions similar to the environment in the human body.

1 is a cross-sectional view of a cell culture apparatus according to an embodiment of the present invention.
2 is a perspective view illustrating a cell culture apparatus according to an embodiment of the present invention.
3A to 3G are cross-sectional views illustrating a method of manufacturing a membrane structure according to an embodiment of the present invention.
FIGS. 4A to 4D are cross-sectional views illustrating pattern shapes of a membrane structure according to an embodiment of the present invention.
5 is a photograph of a membrane according to an embodiment of the present invention observed with a scanning electron microscope.

BRIEF DESCRIPTION OF THE DRAWINGS The advantages and features of the present invention, and how to accomplish them, will become apparent by reference to the embodiments described in detail below with reference to the accompanying drawings. The present invention may, however, be embodied in many different forms and should not be construed as being limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the concept of the invention to those skilled in the art. Is provided to fully convey the scope of the invention to those skilled in the art, and the invention is only defined by the scope of the claims. Like reference numerals refer to like elements throughout the specification.

The terminology used herein is for the purpose of illustrating embodiments and is not intended to be limiting of the present invention. In the present specification, the singular form includes plural forms unless otherwise specified in the specification. As used herein, the terms 'comprises' and / or 'comprising' mean that the stated element, step, operation and / or element does not imply the presence of one or more other elements, steps, operations and / Or additions.

In addition, the embodiments described herein will be described with reference to cross-sectional views and / or plan views, which are ideal illustrations of the present invention. In the drawings, the thicknesses of the films and regions are exaggerated for an effective description of the technical content. Thus, the shape of the illustrations may be modified by manufacturing techniques and / or tolerances. Accordingly, the embodiments of the present invention are not limited to the specific forms shown, but also include changes in the shapes that are generated according to the manufacturing process. For example, the etched area shown at right angles may be rounded or may have a shape with a certain curvature. Thus, the regions illustrated in the figures have schematic attributes, and the shapes of the regions illustrated in the figures are intended to illustrate specific types of regions of the elements and are not intended to limit the scope of the invention.

1 is a cross-sectional view of a cell culture apparatus according to an embodiment of the present invention. 2 is a perspective view illustrating a cell culture apparatus according to an embodiment of the present invention.

Referring to FIGS. 1 and 2, a cell culture apparatus 100 includes a lower structure 10, a membrane structure 20, and a superstructure 30. The lower structure 10 includes a culture liquid storage part 12 and a culture liquid inlet 14. The culture liquid storage part 12 is a concave part of the upper surface of the lower structure 10 and the bottom surface of the culture liquid storage part 12 is located below the upper surface of the lower structure 10. Therefore, the culture medium storage part 12 can be filled with the culture solution used for culturing the cells. The culture medium inlet 14 may have a tube shape and extend from the culture medium reservoir 12 to the side of the lower structure 10.

The substructure 10 may comprise plastic, polymeric materials (e.g., polydimethylsiloxane (PDMS), or glass). The lower structure 10 may be formed by patterning the upper surface of the lower structure 10 so that the culture solution storage part 12 and the culture solution injection part 14 extending to the culture solution storage part 12 . The upper surface of the patterned lower structure 10 may be subjected to an oxygen plasma treatment to be hydrophilically surface-modified.

The membrane structure 20 may be disposed on the lower structure 10. The membrane structure 20 may include a support 21, an anti-etching pattern 23, and a membrane 25. The supporter 21 may be formed to expose the culture medium reservoir 12 on the lower structure 10. The support 21 may include silicon (Si). An anti-oxidation pattern 23 may be formed on the support 21. The anti-oxidation pattern 23 may have a hydrophilic property. The oxidation preventing pattern 23 may include indium tin oxide (ITO), zinc oxide (ZnO), silicon oxide (SiO ₂ ), or titanium dioxide (TiO ₂ ). A membrane 25 may be formed on the anti-oxidation pattern 23. The membrane (25) may have pores (27). The pores (27) may be disposed adjacent to the culture medium storage part (12). The lower surface of the membrane 25 facing the bottom surface of the culture medium reservoir 12 may have hydrophilic properties. The membrane (25) To about 20 < RTI ID = 0.0 >.≪ / RTI > The diameter of the pores 27 is about 0.5? To about 20 < / RTI > The membrane 25 may be formed of a material such as silicon on insulator (SOI), silicon oxide, silicon nitride, a combination of silicon oxide and silicon nitride, a polymer material, glass or metal (e.g., Cr, Ni, Gold (Au)).

The upper structure 30 may be disposed on the membrane structure 20. The upper structure 30 may include a partition wall 31, a cell inlet 33, and a cell outlet 35. The partition 31 may be disposed on the membrane 25. The space partitioned by the partition 31 may be a cell culture unit 37 in which cells are fixed and cultured by the culture solution. One or more barrier ribs 31 may be formed to allow different types of cells to be disposed in each of the cell culture units 37. The cell culture units 37 may be arranged in series or in parallel according to the arrangement of the partitions 31. The cell inlet 33 may be connected to the cell culture unit 37. The cell inlet 33 may be formed according to the number of cells fixed to the cell culture unit 37. The cell outlet (35) may be connected to the cell culture unit (37). Therefore, the cultured cells can be discharged to the cell outlet 35 together with the culture solution.

The cell culture apparatus 100 uses the membrane 25 having a small thickness to immobilize the culture solution provided in the culture solution storage section 12 to the cell culture section 37 located on the culture solution storage section 12 Lt; RTI ID = 0.0 > cells. ≪ / RTI > Therefore, the cells can be supplied with a uniform culture liquid through the membrane 25. In addition, the cell culture apparatus 100 may form a cell discharge port 35 to discharge the waste matter generated from cells in the culture solution to the cell discharge port 35. Therefore, contamination of the culture liquid can be prevented, and the cells can be proliferated under conditions similar to the environment in the human body.

3A through 3G are cross-sectional views illustrating a method of fabricating a microporous membrane structure according to an embodiment of the present invention. 4A to 4D are cross-sectional views illustrating pattern shapes of a microporous membrane structure according to an embodiment of the present invention.

Referring to FIGS. 3A and 3B, a substrate 1 is prepared. The substrate 1 may be a silicon substrate. An etching stopper film (3) is formed on the substrate (1). The etch stop layer 3 may be formed of a material having an etch selectivity with respect to the substrate 1. The etch stop layer 3 may be formed of an oxide film, for example, an indium tin oxide (ITO) film, a zinc oxide film (ZnO), a silicon oxide film (SiO ₂ ), or a titanium dioxide film (TiO ₂ ) . The etch stop layer 3 may be formed of a material having a hydrophilic property. The etch stop layer 3 may be formed by a chemical vapor deposition method, a physical vapor deposition method, or an atomic layer deposition method.

Referring to FIGS. 3C and 3D, a membrane 5 is formed on the etch stop layer 3. The membrane membrane 5 is about 1? To about 20 < RTI ID = 0.0 >. ≪ / RTI > The membrane may be formed by physical vapor deposition (PVD), chemical vapor deposition (CVD), or thermal oxidation. For example, the physical vapor deposition may be an E-beam deposition method or a sputtering deposition method. The chemical vapor deposition method may be performed by, for example, a low pressure CVD method or a plasma- (Plasma Enhanced CVD). The membrane 5 may be formed of a silicon on insulator (SOI) film, a silicon oxide film, a silicon nitride film, a material film in which silicon oxide and silicon nitride are combined, a polymer film, a glass film or a metal film (for example, Film (Ni), gold film (Au)).

The membrane 25 may be formed by patterning the membrane 5. The membrane 25 may arrange a hard mask pattern (not shown) on the membrane 5 to etch a portion of the membrane 5 exposed to the hard mask pattern. Thus, the membrane 25 having a plurality of holes 27 can be formed. The membrane 25 may be formed by performing a dry etching (e.g., reactive ion etching, ion milling) or wet etching on the membrane 5. The wet etching may use, for example, KOH solution or TMAH (Tetramethyl ammonium hydroxide) solution. Referring to FIGS. 4A-4D, the membrane 25 may have various types of patterns. 4A, the side walls 25a of the membrane exposed by the holes 27 may be perpendicular to the etch stop layer. As shown in FIGS. 4B and 4C, the side walls 25a of the membrane may have a concave or tapered shape. 4B, the width of the bottom surface of the holes 27 may be smaller than the width of the tops of the holes 27. When the side walls 25a of the membranes are tapered, The width of the bottom surface of the hole 27 may be larger than the width of the upper portion of the holes 27. The holes 27 may be pores that allow the culture fluid to be absorbed from the culture liquid reservoir 12 (see FIG. 1) into the cell culture section 37 (see FIG. 1) in the membrane structure 20 (see FIG. 1). The holes 27 are about 0.5? To about 20? As shown in FIG. The holes 27 may be formed to have a constant spacing.

Referring to FIG. 3E, the lower surface of the substrate 1 may be etched to form a support 21. The support 21 may be formed by forming an etch mask pattern (not shown) on the lower surface of the substrate 1 and etching a portion of the lower surface of the substrate 1 exposed to the etch mask pattern have. The substrate 1 may be etched by dry etching or wet etching in a direction having anisotropic characteristics. When the dry etching is performed, an ICP-etcher may be used. When the wet etching is performed, TMAH (Tetramethyl ammonium hydroxide) solution may be used. The substrate 1 may be etched until the bottom surface of the etch stop layer 3 is exposed. Since the substrate 1 has an etch selectivity with the etch stop layer 3, the etching of the etch stop layer 3 does not proceed after the etch of the substrate 1 proceeds.

Referring to FIGS. 3F and 3G, the etch stop layer 3 may be patterned or removed to form the membrane structure 20. According to one embodiment, when the etch stop layer 3 is a hydrophilic material, the etch stop layer 3 may be patterned to form the etch stop layer 23. The etch stop pattern 23 may be formed by wet etching or dry etching. The etch stopping pattern 23 may be formed to penetrate the holes 27 of the membrane 35. According to another embodiment, when the etch stop layer 27 is a hydrophobic material as shown in FIG. 3G, the etch stop layer 3 may be removed. The etch stop layer 3 may be removed and the surface of the membrane 25 may be completely exposed. Since the surface of the membrane 25 has a hydrophobic property, surface modification is required to improve the wettability of the surface. A method of modifying the surface of the membrane 25 with a hydrophilic property may be performed by, for example, a method of modifying the surface of the membrane 25 by oxygen plasma treatment or a method of applying a chemical agent containing a surfactant to the membrane 25 ) ≪ / RTI >

The membrane 25 may be formed as an ultra-thin film of a large area using a semiconductor process. Therefore, mass production can be achieved while greatly reducing the manufacturing cost.

Referring again to FIG. 1, a bonding process may be performed in which the lower structure 10 is attached to one surface of the membrane structure 20, and the upper structure 30 is attached to the other surface. Accordingly, the cell culture apparatus 100 including the membrane structure 20 can be formed. The membrane structure 20 may be bonded due to an adhesive or a hot press process.

5 is a photograph of a membrane according to an embodiment of the present invention observed with a scanning electron microscope.

Referring to FIG. 5, the membrane is about 1? Or less and pores having a diameter of about 1 & tilde &

While the present invention has been described in connection with what is presently considered to be practical exemplary embodiments, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, It will be understood. It is therefore to be understood that the above-described embodiments are illustrative and not restrictive in every respect.

10:
12: Culture medium reservoir
14: Culture medium inlet
20: Membrane structure
21: Support
23: anti-oxidation pattern
25: Membrane
27: Voids
31:
33: cell inlet
35: cell outlet
37: Cell culture section
100: Cell culture apparatus

Claims (14)

A lower structure including a culture liquid storage part for storing the culture liquid and a culture liquid injection port connected to the culture liquid storage part and supplying the culture liquid;
A membrane structure including a membrane adjacent to the culture medium reservoir on the lower structure and absorbing the culture medium from the culture medium reservoir through a plurality of pores; And
And an upper structure having a cell injection port for providing cells to the membrane on the membrane structure, and a cell discharge port connected to the cell injection port for discharging the cells. The method according to claim 1,
Wherein the upper structure further comprises a partition disposed on the membrane and partitioning the cell culture section. 3. The method of claim 2,
Wherein the sidewall of the membrane exposed to the partition wall has a vertical, tapered, or concave shape from the lower surface of the membrane. The method according to claim 1,
Wherein the membrane structure comprises: a support disposed on both sides of the culture liquid storage part and supporting the membrane, the membrane structure being disposed on the lower structure; And
Further comprising an etch stop pattern interposed between the support and the membrane. The method according to claim 1,
The membrane is a 1? To 20 < [Lambda] >. The method according to claim 1,
The pores are 0.5? To 20 < [Lambda] >. The method according to claim 1,
Wherein the membrane comprises a silicon on insulator (SOI), silicon oxide, silicon nitride, a combination of silicon oxide and silicon nitride, a polymeric material, a glass or a metal material. Forming an etch stopping film on the semiconductor substrate;
Forming a membrane on the etch stop layer;
Patterning the membrane to form a membrane having holes exposing an upper surface of the etch stop layer;
Etching the lower surface of the semiconductor substrate to form a support for exposing a portion of the etch stopping layer; And
And patterning the etch stopping layer exposed on the support to form an etch stopping pattern passing through the holes. 9. The method of claim 8,
Removing the etch stop pattern to fully expose the surface of the membrane; And
And hydrophilically surface-modifying the surface of the membrane. 10. The method of claim 9,
Wherein the hydrophilic surface modification comprises coating the surface of the membrane with an oxygen plasma treatment on the membrane surface or a chemical solvent containing a surfactant. 9. The method of claim 8,
Wherein the etch stop layer is formed of a material having etch selectivity with respect to the semiconductor substrate. 9. The method of claim 8,
Wherein the etch stop layer is formed of a material having hydrophilicity. 9. The method of claim 8,
The membrane membrane is a 1? To 20 < RTI ID = 0.0 >.≪ / RTI > 9. The method of claim 8,
Wherein the membrane is formed by performing a wet etching or a dry etching process.

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