JP5790056B2 - Cell culture vessel - Google Patents

Description

  The present invention relates to a cell culture container excellent in antifogging property and anti-contamination property when taken out from a culture thermostat.

In order to culture cells efficiently, it is necessary to culture them under conditions suitable for growth. For this reason, cells are usually cultured in a culture constant temperature bath that can be maintained at a predetermined temperature and humidity.
In addition, when observing the state of cells in culture, it is necessary to take out the cell culture container containing the cells from the thermostat. In order to prevent contamination when the cells are taken out from the thermostat, cell culture is usually performed. A cell culture container with a lid is used as the container.
Here, for example, a temperature condition of about 37 ° C. is used for cells such as humans, but if a cell culture vessel placed for a long time under such a temperature condition is taken out of the thermostat and placed under a room temperature condition, Water vapor contained in the culture container is condensed on the inner surface of the lid covering the cell culture container, and a large amount of water droplets are attached. As a result, there is a problem that it becomes difficult to observe cells in a state covered with a lid.

With respect to such problems, water having a wiper on the inner surface of the lid (Patent Document 1), water having a hydrophilic polymer layer on the inner surface of the lid, and further adhering to the hydrophilic polymer layer on the outer periphery of the lid The thing (patent document 2) etc. which have a collar part to receive are disclosed. According to such a method, the water condensed on the inner surface of the lid can be prevented from adhering in the form of water droplets, and the cells can be observed with high visibility.
However, the thing which provided the wiper in the inner surface of the lid | cover like patent document 1 had malfunctions, such as a reduction of the observable area of a culture container and an operator's effort.
Further, Patent Document 2 is characterized in that condensed water accumulates in a water receiving portion provided in the lid portion. However, such a structure has poor handling properties compared to an original culture vessel, and medium replacement. There has been a problem that the risk of contamination increases due to the outflow and scattering of water accumulated in the experimental operation.

JP-A-5-184349 JP 2010-51200 A

  The present invention has been made in view of the above problems, and a main object of the present invention is to provide a cell culture container excellent in anti-fogging property and anti-contamination property when taken out from a culture constant temperature bath.

  In order to achieve the above object, the present invention provides a cell culture container having a cell culture part having a recess surrounded by a side wall and a lid part covering the cell culture part, The inner surface is formed so as to surround a part of the opening of the concave portion of the cell culture portion so as to overlap in plan view, and the upper surface of the side wall. And a water-blocking portion formed so as to overlap with each other in plan view.

According to the present invention, the water film forming portion is formed on the inner surface of the lid portion so as to overlap with a part of the opening of the concave portion in plan view. In addition, it is possible to prevent adhesion of water droplets and to exhibit excellent antifogging properties.
Moreover, contamination can be stably prevented by having the water blocking part formed on the inner surface of the lid part so as to overlap the upper surface of the side wall in plan view.

  In the present invention, the water film forming part is preferably a hydrophilic part or a fine uneven part. This is because adhesion of water droplets to the water film forming portion can be prevented, and a water film can be stably formed.

  In the present invention, the water blocking part preferably includes a hydrophobic part, a water absorbing part, and / or a step part. By including such a part, it is possible to stably prevent water adhering to the water film forming part from flowing out of the cell culture part.

  INDUSTRIAL APPLICATION This invention has an effect that the cell culture container excellent in the anti-fogging property at the time of taking out from the culture thermostat and the anti-contamination property can be provided.

It is a schematic sectional drawing which shows an example of the cell culture container of this invention. It is the XX sectional view taken on the line of FIG. It is a schematic plan view which shows the other example of the cell culture container of this invention. It is explanatory drawing explaining the cell culture part in this invention. It is explanatory drawing explaining the cell culture part in this invention. It is explanatory drawing explaining the side wall in this invention. It is explanatory drawing explaining the side wall in this invention. It is a schematic sectional drawing which shows the other example of the cell culture container of this invention. It is a schematic sectional drawing which shows the other example of the cell culture container of this invention. It is a schematic sectional drawing which shows the other example of the cell culture container of this invention. It is a schematic sectional drawing which shows the other example of the cell culture container of this invention. It is a schematic sectional drawing which shows the other example of the cell culture container of this invention. It is a schematic sectional drawing which shows the other example of the cell culture container of this invention.

The present invention relates to a cell culture container.
Hereinafter, the cell culture container of the present invention will be described in detail.

  The cell culture container of the present invention is a cell culture container having a cell culture part having a recess surrounded by a side wall and a lid part covering the cell culture part, and the inner surface of the lid part has A water film forming part formed so as to overlap a part of the opening of the cell culture part in plan view, and so as to surround the water film forming part, and so as to overlap the upper surface of the side wall in plan view It has the formed water interruption | blocking part, It is characterized by the above-mentioned.

  Such a cell culture container of the present invention will be described with reference to the drawings. FIG. 1 is a schematic cross-sectional view showing an example of the cell culture container of the present invention. As illustrated in FIG. 1, the cell culture container 20 of the present invention has a cell culture part 3 having a recess 2 surrounded by a side wall 1 and a lid part 10 covering the cell culture part 3. It is. The lid portion 10 mainly comprises a lid base material 13, and a water film is formed on the inner surface of the lid base material 13 so as to overlap a part of the opening of the concave portion 2 of the cell culture part 3 in plan view. The water blocking part 12 is formed so as to surround the part 11 and the water film forming part 11 and is formed so as to overlap the upper surface of the side wall 1 in plan view.

According to the present invention, the water film forming part is formed on the inner surface of the lid part so as to overlap with a part of the opening in plan view, so that when the water film is removed from the culture thermostat, water droplets are formed on the lid inner surface. Can be prevented and a water film can be formed. As a result, excellent anti-fogging properties can be exhibited in such a water film forming portion, and it can be used as an observation region for favorably observing cells in culture in the recess.
Further, by having a water blocking part formed on the inner surface of the lid part so as to overlap the upper surface of the side wall in plan view, water attached to the water film forming part flows out of the cell culture part in plan view. Can be prevented. For this reason, it can prevent that the exterior of a cell culture container and the culture solution in a cell culture part contact through the water adhering to the lid | cover by dew condensation. As a result, contamination can be stably prevented.
On the other hand, when the water film forming part is arranged wider than the opening of the concave part of the cell culture part in plan view, the dew condensation water adhering to the water film forming part is, for example, the side wall of the cell culture part. May then flow down to the outside of the cell culture vessel along the upper surface of the side wall. Then, the continuous water marks formed by the movement of water from the inside of the cell culture section to the outside of the cell culture container are connected to the inside of the recess of the cell culture section and the outside of the cell culture container. This increases the possibility of contamination in the recess of the cell culture part.

The cell culture container of the present invention has at least a cell culture part and a lid part.
Hereafter, each structure of the cell culture container of this invention is demonstrated in detail.

1. Cell culture part The cell culture part used for this invention has a recessed part with the circumference | surroundings surrounded by the side wall. In addition, the cells and the medium are held in the recesses in the present invention, and the cells are cultured.

Such a cell culture part is not particularly limited as long as it has at least one recess and can be covered with a lid part. As such a cell culture part, as long as it is integrally formed, it may have two or more recesses. For example, the petri dish as shown in FIG. 1 already described, FIG. 2 and FIG. It can be made to be the same as that of a general cell culture container, such as a multi-well plate and cell culture bottles, such as a culture flask.
2 is a cross-sectional view taken along the line XX of FIG. 3, and FIG. 3 is a schematic plan view showing another example of the cell culture container. Moreover, about the code | symbol in FIGS. 2-3, since the same member as the thing in FIG. 1 is shown, description here is abbreviate | omitted.

The material constituting the cell culture section in the present invention is not particularly limited as long as cell culture and observation can be easily performed, and can be the same as a general cell culture vessel. .
Specifically, polyethylene terephthalate (PET), polystyrene (PS), polycarbonate (PC), TAC (triacetyl cellulose), polyimide (PI), nylon (Ny), low density polyethylene (LDPE), medium density polyethylene (MDPE) ), Acrylic materials such as vinyl chloride, vinylidene chloride, polyphenylene sulfide, polyether sulfone, polyethylene naphthalate, polypropylene, urethane acrylate, cellulose, glass and the like. In addition, a resin such as a biodegradable polymer such as polylactic acid, polyglycolic acid, polycaprolactan, or a copolymer thereof can be used.
In the present invention, among these, polyethylene terephthalate, polystyrene, and polycarbonate can be preferably used, and in particular, polystyrene can be preferably used. This is because the cytotoxicity is low.

As long as cells can be stored and cultured in the present invention, the shape and volume thereof are not limited, and can be the same as those used for general cell culture. For example, as shown in FIG. 1 to FIG. 3 described above, the columnar shape in which the side wall is erected vertically, or the truncated cone shape and the conical shape in which the side wall is inclined as illustrated in FIG. A polygonal pyramid shape, a polygonal frustum shape, or a curved side wall as exemplified in FIG. 5 can be used.
The reference numerals in FIGS. 4 to 5 indicate the same members as those in FIG. Moreover, the site | part shown by A in FIG. 4 and FIG. 5 shows the upper surface of a side wall.

As for the surface of the recess in the present invention, the material constituting the cell culture part may be exposed, or it may be subjected to a surface treatment if necessary.
Examples of such surface treatment include ultraviolet irradiation, corona treatment, plasma treatment, and application of a coating solution containing collagen, fibronectin, laminin, poly L-lysine, and the like.

Note that the opening of the concave portion of the cell culture portion in the present invention overlaps with the water film forming portion in plan view, and the opening portion of the concave portion of the cell culture portion referred to here is the upper side (lid) of the concave portion. The opening region on the side facing the part) refers to a region surrounded by the upper surface of the side wall.
In addition, the upper surface of the side wall of the cell culture unit in the present invention overlaps with the water blocking unit in plan view, and the upper surface of the side wall here is continuous with the upper surface of the side wall as well as the upper surface of the side wall. And the upper surface of the site | part which has the upper surface of comparable height is also included. Here, the same level of height indicates that the surface includes an uneven shape on the upper surface in the present invention. Specifically, as shown in FIG. 2 described above, when a concave portion is formed on a flat plate-like member, other than the portion where the concave portion is formed on the surface of the plate-like member. Is the upper surface of the side wall. Further, when the cell culture part has a shape such that a plurality of recesses are connected on the opening side as illustrated in FIG. 6, or a plurality of recesses are connected on the bottom side as illustrated in FIG. In the case of such a shape, the part indicated by A is the upper surface of the side wall. Furthermore, as shown in FIGS. 4 to 5 already described, in the case where the side wall of the cell culture part is inclined or the side wall is curved, the part indicated by A and facing the lid part is respectively provided. It becomes the upper surface of the side wall.
6 to 7 indicate the same members as those in FIG. 1, and thus the description thereof is omitted here.

2. Lid part The cover part in this invention covers the said cell culture part, and has a water film formation part and a water interruption | blocking part in an inner surface.

(1) Water film formation part The water film formation part in this invention is formed so that it may overlap with a part of opening part of the said cell culture part on planar view, and forms a water film with the water adhering by dew condensation. It is a part to do.

  The formation location of the water film forming portion in the present invention is not particularly limited as long as it is formed so as to overlap with a part of the opening of the cell culture portion in plan view. It is preferable that they are overlapping portions. This is because the cells cultured in the recesses of the cell culture part can be observed with good visibility.

  In the present invention, the water film forming part is preferably formed with an area of 80% or more of the area of the opening, and in particular, the water film forming part is formed with an area within the range of 90% to 98%. It is preferable. This is because, when the water film forming part is formed with the above area, the cells cultured in the concave part of the cell culture part can be observed with high visibility. Moreover, it is because the water on a water film formation part can prevent moving outside a recessed part.

  Such a water film forming portion is not particularly limited as long as it can stably form a water film, and examples thereof include a hydrophilic portion and a fine uneven portion.

(A) Hydrophilic part The hydrophilic part in this invention has hydrophilicity, and the condensed water which adhered can form a water film.
In the present invention, the water contact angle on the surface is preferably 50 ° or less, and particularly preferably 30 ° or less. This is because the water film can be stably formed by the water contact angle.
The water contact angle refers to the contact angle with water or a liquid having an equivalent contact angle. After a predetermined time has elapsed after a drop (a certain amount) of pure water has been dropped on the surface of the hydrophilic portion. The water droplet shape can be observed using a microscope or a CCD camera, and can be measured using a method for physically obtaining the contact angle.
More specifically, using a contact angle measuring device (CA-Z type manufactured by Kyowa Interface Science Co., Ltd.) (measured 30 seconds after dropping a drop of pure water from a microsyringe), from the result, or The result can be obtained as a graph.
In addition, the lower limit of the water contact angle is not particularly limited because the smaller the effect, the more effective it can be set according to the formability of the hydrophilic part, the cost of the constituent material, and the like. it can.

As a material constituting such a hydrophilic portion, any material may be used as long as it has a transparency that allows the cells in the cell culture portion to be observed from above the lid portion and has a hydrophilic property that can stably form a water film. Although not limited, for example, resin materials such as polystyrene, polyethylene terephthalate, and polycarbonate subjected to a predetermined surface treatment, glass, and the like, furthermore, inorganic materials such as silicon oxide, orientation groups of self-assembled monolayers In which is substituted with a hydroxyl group.
In the present invention, among them, polystyrene and polyethylene terephthalate subjected to a predetermined surface treatment can be preferably used. This is because by using such a material, a hydrophilic portion having high hydrophilicity can be easily formed.
Specific examples of the surface treatment applied to the resin material include application of an antifogging agent such as a surfactant, irradiation with plasma, corona, microwave, electron beam, ultraviolet light, and the like. Can do.
Specific examples of the resin material subjected to such a predetermined surface treatment include a polystyrene sheet (Sandick sheet manufactured by Sandic Corporation) in which an antifogging agent is applied to a sheet made of polystyrene. Can do.

  Here, when the material is silicon oxide, that is, when the hydrophilic portion is a silicon oxide film, the ratio of oxygen atoms contained in the silicon oxide film is 150 to 300, more particularly, 100 to 100 Si atoms. It is preferable to be within the range of 180 to 250.

  Further, the ratio of carbon atoms contained in the silicon oxide film is preferably 50 or less, more preferably 20 or less, with respect to 100 Si atoms.

  When the ratio of oxygen atoms in the silicon oxide film is within the above range, the silicon oxide film can be a high-purity silicon oxide film with few impurities such as carbon atoms and fluorine atoms. Further, when the ratio of carbon atoms exhibiting hydrophobicity in the silicon oxide film is within the above range, a higher hydrophilic silicon oxide film can be obtained.

  Here, the component ratio in the silicon oxide film in the present invention is a value measured by XPS (X-ray Photoelectron Spectroscopy). The analysis method by XPS will be described below. When a solid surface is irradiated with X-rays in a vacuum, electrons are scattered from the surface atoms given energy by the X-rays. Since these electrons are generated by irradiation with light such as X-rays, they are called photoelectrons. Since these photoelectrons have energy inherent to the elements, it is possible to perform qualitative analysis and quantitative analysis of elements by measuring the energy distribution. Become. In addition, photoelectrons generated deep from the surface lose their energy before coming out to the surface and are difficult to measure. The escape depth of electrons having a kinetic energy of 1000 eV is several nm (tens of atomic layers). Therefore, it is possible to obtain information on the outermost surface. Furthermore, in order to measure the deep part, the surface needs to be sputtered with ions such as argon, and selective sputtering occurs depending on the type of element, so correction is required for quantification.

In addition, the self-assembled monomolecular film in the case where the above-described material is obtained by replacing the orientation group of the self-assembled monomolecular film with a hydroxyl group is a solid / liquid or solid / gas interface, and organic molecules spontaneously It is an organic thin film that spontaneously forms a monomolecular film while gathering together to form an aggregate. For example, when a substrate made of a specific material is exposed to a solution or vapor of an organic molecule having a high chemical affinity with the substrate material, the organic molecule chemically reacts and adsorbs on the substrate surface. If the organic molecule consists of two parts, a functional group with high chemical affinity and an alkyl group that does not cause any chemical reaction with the substrate, the molecule reacts when the functional group with high affinity is at its end. The functional group having a high chemical affinity at the sex end faces the substrate side and the alkyl group is adsorbed outward. When alkyl groups gather together, the whole becomes stable, so organic molecules gather spontaneously during the process of chemisorption. Since molecular adsorption requires a chemical reaction between the substrate and the terminal functional group, once the surface of the substrate is covered with organic molecules to form a monomolecular film, molecular adsorption does not occur thereafter. Absent. As a result, an organic monomolecular film in which molecules are densely gathered and aligned is formed. In the present invention, such a film is a self-assembled monolayer. Here, the reactive terminal group bonded to the substrate is an adsorbing group, and the group oriented to the outside is an aligning group.
Accordingly, the substitution of the orientation group of the self-assembled monolayer in the present invention with a hydroxyl group means that the orientation group is substituted with a hydroxyl group after the formation of the self-assembled monolayer. .
In addition, as a method of substituting the orientation group with a hydroxyl group, a method of irradiating with high energy in a reactive atmosphere can be mentioned. In particular, a method of plasma irradiation in an atmosphere containing oxygen gas or ultraviolet irradiation in an oxygen-containing atmosphere is desirable.

  Here, the self-assembled monolayer is preferably formed using a self-assembled monolayer forming material represented by the following general formula (1) as a raw material.

R ¹ _α XR ² _β (1)

Here, R ¹ is an alkyl group having 1 to 30 carbon atoms or an aryl group (benzene ring), and the carbon group includes those partially having a branched chain or multiple bonds. In addition, examples of the element bonded to carbon include halogen such as fluorine and chlorine, hydrogen, nitrogen, and the like. R ² is halogen, or —OR ³ (R ³ is an alkyl group having 1 to 6 carbon atoms, an aryl group, or an allyl group. Note that carbon is not only oxygen or hydrogen, but also halogen and nitrogen. A bonded group is also included). X is one element selected from the group consisting of Si, Ti, Al, C and S. Here, α and β are integers of 1 or more, and α + β is 2 to 4.

Here, R ¹ is the above-mentioned orientation group, R ² is the above-mentioned adsorption group, and X is a substance that becomes the nucleus of the self-assembled monolayer. The substance having the above structure forms a self-assembled monolayer.

  Specific examples of the self-assembled monolayer forming material shown above include octadecyltrimethoxysilane, octadecyltriethoxysilane, octadecyltrichlorosilane, octadecylmethyldiethoxysilane, octadecyldimethylmethoxysilane, octadecylmethyldimethoxysilane, octadecyl Methoxydichlorosilane, octadecylmethyldichlorosilane, octadecyldimethyl (dimethylamino) silane, octadecyldimethylchlorosilane, nonylchlorosilane, octenyltrichlorosilane, octenyltrimethoxysilane, octylmethyldichlorosilane, octylmethyldiethoxysilane, octyltrichlorosilane, Octyltriethoxysilane, Octyltrimethoxysilane, Pentafluorophenylpropyltri Lolosilane, pentafluorophenylpropyltrimethoxysilane, pentyltriethoxysilane, pentyltrichlorosilane, phenethyltrichlorosilane, phenethyltrimethoxysilane, phenyldichlorosilane, phenyldiethoxysilane, phenylethyldichlorosilane, phenylmethyldimethoxysilane, phenyltrimethoxysilane , Phenyltrichlorosilane, phenyltriethoxysilane, propyltriethoxysilane, propyltrimethoxysilane, propyltrichlorosilane, tetradecyltrichlorosilane, (3,3,3-trifluoropropyl) dimethylchlorosilane, (3,3,3- (Trifluoropropyl) trichlorosilane, (3,3,3-trifluoropropyl) trimethoxysilane, (3,3,3-toe) Fluoropropyl) triethoxysilane, 3,3,4,4,5,5,6,6,6-nonafluorohexylmethyldichlorosilane, 3,3,4,4,5,5,6,6,6- Nonafluorohexyltrichlorosilane, isooctyltrimethoxysilane, isobutylmethyldichlorosilane, isobutyltriethoxysilane, isobutyltrichlorosilane, hexyltrichlorosilane, hexyltriethoxysilane, hexyltrimethoxysilane, hexenyltrichlorosilane, hexyldichlorosilane, hexadecyl Trichlorosilane, hexadecyltrimethoxysilane, hexadecyltriethoxysilane, heptyltrichlorosilane, (heptadecafluoro-1,1,2,2-tetrahydrodecyl) triethoxysilane, (heptadecafluoro- 1,1,2,2-tetrahydrodecyl) trichlorosilane, (heptadecafluoro-1,1,2,2-tetrahydrodecyl) methyldichlorosilane, (heptadecafluoro-1,1,2,2-tetrahydrodecyl) Dimethylchlorosilane, eicosyltrichlorosilane, dodecyltrimethoxysilane, dodecyltriethoxysilane, dodecyltrichlorosilane, dodecylmethyldichlorosilane, dodecyldimethylchlorosilane, diphenyldiethoxysilane, diphenyldichlorosilane, diphenyldiethoxysilane, diphenyldimethoxysilane, dimethoxy Methyl-3,3,3-trifluoropropylsilane, decylmethyldichlorosilane, decyltrichlorosilane, decyltriethoxysilane, decyltrimethoxysilane, decyldimethyl L-chlorosilane, cyclohexylmethyltrichlorosilane, cyclohexyltrichlorosilane, 3-chloropropylmethyldimethoxysilane, 3-chloropropyltrimethoxysilane, 3-chloropropyltriethoxysilane, 3-chloropropyltrichlorosilane, chlorophenyltrichlorosilane, butyltrimethoxy Examples thereof include silane and butyltrichlorosilane.

  In the present invention, among them, octadecyltrimethoxysilane, octadecyltriethoxysilane, octadecyltrichlorosilane, nonylchlorosilane, octenyltrichlorosilane, octenyltrimethoxysilane, octyltrichlorosilane, octyltriethoxysilane, octyltrimethoxysilane, pentafluoro Phenylpropyltrichlorosilane, pentafluorophenylpropyltrimethoxysilane, pentyltriethoxysilane, pentyltrichlorosilane, phenethyltrichlorosilane, phenethyltrimethoxysilane, phenyltrimethoxysilane, phenyltrichlorosilane, phenyltriethoxysilane, propyltriethoxysilane, propyltri Methoxysilane, propyltrichlorosilane, tetradecylto Chlorosilane, (3,3,3-trifluoropropyl) trichlorosilane, (3,3,3-trifluoropropyl) trimethoxysilane, (3,3,3-trifluoropropyl) triethoxysilane, 3,3, 4,4,5,5,6,6,6-nonafluorohexyltrichlorosilane, isooctyltrimethoxysilane, isobutyltriethoxysilane, isobutyltrichlorosilane, hexyltrichlorosilane, hexyltriethoxysilane, hexyltrimethoxysilane, hexenyl Trichlorosilane, hexadecyltrichlorosilane, hexadecyltrimethoxysilane, hexadecyltriethoxysilane, heptyltrichlorosilane, (heptadecafluoro-1,1,2,2-tetrahydrodecyl) triethoxysilane, (heptadeca (Luoro-1,1,2,2-tetrahydrodecyl) trichlorosilane, eicosyltrichlorosilane, dodecyltrimethoxysilane, dodecyltriethoxysilane, dodecyltrichlorosilane, dimethoxymethyl-3,3,3-trifluoropropylsilane, decyl Trichlorosilane, decyltriethoxysilane, decyltrimethoxysilane, decyldimethylchlorosilane, cyclohexylmethyltrichlorosilane, cyclohexyltrichlorosilane, 3-chloropropyltrimethoxysilane, 3-chloropropyltriethoxysilane, 3-chloropropyltrichlorosilane, chlorophenyl Trichlorosilane, butyltrimethoxysilane, and butyltrichlorosilane are preferred, especially octadecyltrimethoxysilane and octadecyltriethoxy. Sisilane, octadecyltrichlorosilane, octyltriethoxysilane, phenyltriethoxysilane, (tridecafluoro-1,1,2,2-tetrahydrooctyl) triethoxysilane, isobutyltrimethoxysilane, (heptadecafluoro-1,1, 2,2-tetrahydrodecyl) triethoxysilane, dodecyltriethoxysilane, and 3-chloropropyltrimethoxysilane are preferred.

  The thickness of the hydrophilic portion in the present invention is not particularly limited as long as desired hydrophilicity can be exhibited, and can be appropriately set depending on the material and the like, for example, 1 nm to 1 mm. Can be about. More specifically, when the material constituting the hydrophilic portion is an inorganic material such as silicon oxide or a material in which the orientation group of the self-assembled monolayer is substituted with a hydroxyl group, the range of 1 nm to 150 nm. In particular, it is preferably 1 nm to 100 nm, and particularly preferably in the range of 1 nm to 50 nm. This is because the hydrophilic portion can be formed stably. Moreover, when a hydrophilic part is formed by affixing the sheet | seat which consists of resin materials, such as a polystyrene, a polyethylene terephthalate, a polycarbonate, to a cover base material, it is preferable that it is about 100 micrometers-1 mm. This is because pasting is easy.

The method for forming the hydrophilic portion in the present invention is not particularly limited as long as it is a method capable of stably forming a hydrophilic portion having a desired hydrophilicity. A method of forming a hydrophilic part by applying and drying a coating liquid containing the coating liquid, a method of attaching the hydrophilic part to a lid substrate, and the like can be used.
Further, when the material is an inorganic material such as silicon oxide, a method using a vapor deposition method or the like may be used.

(B) Fine uneven part The fine uneven part in this invention becomes a film | membrane form of the water adhering by the fine unevenness | corrugation of the surface.
The fine irregularities of such fine irregularities are not particularly limited as long as a water film can be formed, and even if a water film is formed, it has transparency. For example, the arithmetic average roughness Ra is preferably in the range of 5 nm to 200 nm, more preferably in the range of 5 nm to 100 nm, and particularly preferably in the range of 5 nm to 50 nm. This is because the water film can be stably formed when the surface roughness of the fine irregularities is within the above range.
The arithmetic average roughness Ra is defined by JISB 0601.
More specifically, the surface roughness can be obtained by measuring in a tapping mode using an atomic force microscope (manufactured by Seiko Instruments Inc .; SPI3800N).

  The material constituting the fine irregularities in the present invention is not particularly limited as long as it has transparency and can form desired fine irregularities on the surface. However, high energy is applied in a reactive atmosphere. It is preferable that the material is formed with fine irregularities on the surface by irradiation, and in particular, a material capable of forming fine irregularities on the surface by irradiation with plasma or ultraviolet light is preferable. This is because fine irregularities can be obtained by plasma irradiation or ultraviolet light irradiation, which is preferable in terms of manufacturing efficiency and cost.

  Examples of such materials include polyolefin (PO) resins such as homopolymers or copolymers or copolymers such as ethylene, polypropylene, and butene, amorphous polyolefin resins (APO) such as cyclic polyolefins, Polyester resins such as polyethylene terephthalate (PET) and polyethylene 2,6-naphthalate (PEN), polyamide (PA) resins such as nylon 6, nylon 12, copolymer nylon, polyvinyl alcohol (PVA) resin, ethylene-vinyl Polyvinyl alcohol resins such as alcohol copolymers (EVOH), polyimide (PI) resins, polyetherimide (PEI) resins, polysulfone (PS) resins, polyethersulfone (PES) resins, polyether ethers Ketone (PEEK) resin Carbonate (PC) resin, polyvinyl butyrate (PVB) resin, polyarylate (PAR) resin, polystyrene resin, ethylene-tetrafluoroethylene copolymer (ETFE), ethylene trifluoride chloride (PFA), Tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (FEP), vinylidene fluoride (PVDF), vinyl fluoride (PVF), perfluoroethylene-perfluoropropylene-perfluorovinyl ether-copolymer (EPA), etc. A resin material such as a fluorine-based resin can be used.

  In addition to the resin materials listed above, a resin composition comprising an acrylate compound having a radical-reactive unsaturated compound, a resin composition comprising the acrylate compound and a mercapto compound having a thiol group, epoxy acrylate, urethane acrylate It is also possible to use a photocurable resin such as a resin composition in which an oligomer such as polyester acrylate or polyether acrylate is dissolved in a polyfunctional acrylate monomer, and a mixture thereof. Furthermore, it is also possible to use what laminated | stacked 1 or 2 or more types of these resin by means, such as a lamination and a coating, as a base material. Furthermore, a base material obtained by coating these resins on a solid surface such as glass or a silicon wafer may be used, or an inorganic material such as glass or silicon may be used. Further, a material constituting the hydrophilic portion as described above may be used.

  In the present invention, among them, a polyester resin, a polystyrene resin or a polycarbonate resin is preferable. Among polyester resins, polyethylene terephthalate and polyethylene naphthalate are particularly preferable. This is because, when the material is any of these substances, it is easy to form fine irregularities on the surface, and processing is facilitated.

  In the present invention, when the material is a resin material, the fine uneven portion may be unstretched, but is particularly stretched, that is, a layer made of the material (hereinafter referred to as fine unevenness). It may be a part forming layer.) Is preferably formed by forming fine irregularities on the stretched layer. By stretching the layer for forming fine irregularities, regular orientation becomes possible, and fine irregularities are easily formed regularly on the entire surface, and the water film can be more stably formed by the fine irregularities. Because it becomes possible. In addition, even when unstretched, random fine irregularities are formed, so that a water film can be stably formed.

  As a specific stretching method, a known method such as uniaxial stretching, tenter-type sequential biaxial stretching, tenter-type simultaneous biaxial stretching, tubular simultaneous biaxial stretching, and the like for an unstretched fine uneven portion forming layer, The film can be stretched in the flow (vertical axis) direction of the fine uneven portion forming layer or in the direction perpendicular to the flow direction (horizontal axis) of the fine uneven portion forming layer. The draw ratio in this case can be appropriately selected according to the material used as the raw material for the fine uneven portion forming layer, but is preferably 2 to 10 times in each of the vertical axis direction and the horizontal axis direction.

The method for forming fine irregularities in the present invention is not particularly limited as long as the method can accurately form fine irregularities having fine irregularities of a desired size. For example, fine irregularities are formed on a lid substrate. A method for forming fine irregularities after forming the layer for forming fine irregularities before forming, a method for attaching the fine irregularities to the substrate after forming the fine irregularities, and the like can be used.
In addition, about the formation method of the fine uneven | corrugated | grooved part formation layer, it can be made to be the same as that of the formation method of a hydrophilic part.

  The method for forming fine irregularities in the present invention is not particularly limited as long as it is a method capable of forming fine irregularities of a desired size on the surface of the layer for forming fine irregularities made of the above materials. It is also possible to use solution processing such as forming micropores in the development and the like, but in the present invention, it is preferable to irradiate high energy in a reactive atmosphere. As a result, the gas in the reactive atmosphere is excited to be in a high energy state and reacts with a specific portion of the surface of the fine uneven portion forming layer, thereby easily making fine unevenness on the surface of the fine uneven portion forming layer surface. It is because it becomes possible to form in this.

Here, the reactive atmosphere in the present invention is an atmosphere containing a substance that is activated by the high-energy irradiation, and differs depending on the method of high-energy irradiation. An atmosphere of a gas such as O ₂ , N ₂ O, NO, NO ₂ , CO ₂ , CO, H ₂ , He, N ₂ , Ar, and the like can be given, and among them, a gas containing an oxygen atom is preferable.

  Specific examples of the high energy irradiation method include plasma irradiation, ion irradiation, radical irradiation, light irradiation, and the like. Among these, plasma irradiation and ultraviolet light irradiation are preferable.

  As the method for forming the fine unevenness in the present invention, among the above, a plasma treatment using a gas containing oxygen atoms and irradiating plasma or a dry treatment irradiating vacuum ultraviolet light in an oxygen-containing atmosphere is used. preferable. As a result, it is possible to introduce a hydrophilic group to the surface of the fine uneven portion forming layer at the same time as forming the fine uneven portion on the surface of the fine uneven portion forming layer, thereby further improving the formation stability of the water film. It is because it can be made.

(2) Water blocking part The water blocking part in the present invention is formed so as to surround the water film forming part and is formed so as to overlap the upper surface of the side wall in plan view. It is a site | part which prevents the water adhering to moving to the exterior of the cell culture container of this invention.
Here, overlapping with the upper surface of the side wall in plan view means to cover substantially all of the upper surface of the side wall, and there is a case where a part of the upper surface is not covered within a range not hindering the effect of the present invention. Is included. In the present invention, it is preferable that 90% or more of the area of the upper surface of the side wall is covered, and it is preferable that 95% or more is covered, particularly 100%, that is, all are preferably covered.
Therefore, normally, as shown in FIG. 8, a water blocking part is formed so as to cover at least all of the A region.
In addition, about the code | symbol in FIG. 8, since it shows the same member as the thing in FIG. 1, description here is abbreviate | omitted.

In the present invention, the water blocking portion is not particularly limited as long as it is formed so as to surround the water film forming portion and overlap with the upper surface of the side wall in plan view. However, it may or may not be adjacent to the water film forming part, but as shown in FIG. 9, it includes a portion that overlaps with the portion formed continuously with the upper surface of the side wall in plan view. In particular, it is preferable to include a portion overlapping in plan view with all of the portions other than the opening, particularly preferably including a portion overlapping in plan view with a part of the opening. It is preferable to be all inner surfaces other than the region where the water film forming portion is formed. This is because the water blocking portion is formed at the above-mentioned formation place, thereby preventing water from moving more stably.
Note that the reference numerals in FIG. 9 indicate the same members as those in FIG. 1, and thus the description thereof is omitted here.

  Such a water blocking part is not particularly limited as long as it can stably prevent the movement of water. For example, the water blocking part includes a hydrophobic part, a water absorbing part, a step part, and the like. Can be mentioned.

(A) Hydrophobic part The hydrophobic part in the present invention has hydrophobicity to prevent the dew condensation adhering to the water film forming part from moving outside the cell culture container.
In the present invention, the water contact angle on the surface is preferably 60 ° or more, and particularly preferably 80 ° or more. This is because the water contact angle can prevent the movement of water stably.
The upper limit of the water contact angle is not particularly limited because the effect can be exhibited as the value increases, and the upper limit of the water contact angle is set as appropriate depending on the formability of the hydrophobic portion, the cost of the constituent material, and the like. can do.

  The material constituting the hydrophobic part in the present invention is not particularly limited as long as it has hydrophobicity that can stably prevent water movement, and may be transparent or opaque. Although it is good, the material which comprises a lid | cover base material which is mentioned later can be used. That is, the exposed part of the lid in a general cell culture container can be used as the hydrophobic part. Specifically, it can be the same as the material constituting the cell culture section.

  The thickness of the hydrophobic portion in the present invention is not particularly limited as long as the hydrophobic portion can exhibit a desired hydrophobicity. For example, it can be the same as the hydrophilic portion.

  The method for forming the hydrophobic portion in the present invention is not particularly limited as long as it can stably form a hydrophobic portion having a desired hydrophobicity, but the same method as that for the hydrophilic portion is used. be able to.

(B) Water absorption part The water absorption part in this invention prevents the movement to the exterior of the cell culture container of the condensed water adhering to a water film formation part by absorbing the water which has moved from the water film formation part. Is.
Such a water-absorbing part is not particularly limited as long as it has water absorption, but the water absorption per 1 m ³ (g / m ³ ) is in the range of 1-60. Among them, it is preferable that it is in the range of 5-30. This is because the water absorption can be stably prevented by having the water absorption property.
The amount of water absorption is determined by measuring the mass after being dried in a desiccator and measuring the mass after being immersed in water at 4 ° C. for 1 hour.

The material constituting the water absorbing portion in the present invention is not particularly limited as long as it can absorb a desired amount of water, for example, a gel such as sodium polyacrylate, polyethylene glycol, Examples thereof include porous materials such as polyurethane, and fiber materials such as polyester.
In the present invention, sodium polyacrylate is particularly preferable. This is because a water-absorbing portion having excellent water absorption can be easily formed by using the above material.

  The thickness of the water-absorbing part in the present invention is not particularly limited as long as the water-absorbing part can exhibit a desired water absorption property. For example, it can be the same as the hydrophilic part.

  The method for forming the water-absorbing part in the present invention is not particularly limited as long as it can stably form a water-absorbing part having a desired water-absorbing property, but the same method as the hydrophilic part is used. be able to.

(C) Stepped portion In the present invention, the stepped portion prevents movement of condensed water adhering to the water film forming portion to the outside by blocking the water moved from the water film forming portion by the step. It is.
Such a stepped portion is not particularly limited as long as the thickness is different from that of the adjacent opening side member, but the difference in thickness between the adjacent opening side member is 5 μm or more. In particular, the thickness is preferably 10 μm or more. This is because the movement of water to the outside can be stably prevented when the difference in thickness is within the above range.
Here, as a specific example in which the thickness is different from the adjacent opening side member, when the stepped portion as the water blocking portion is adjacent to the water film forming portion on the opening side, the thickness is different from the thickness of the water film forming portion. When there is another water blocking part such as a hydrophobic part between the water film forming part and the water film forming part, the water blocking part is different from the thickness of the other water blocking part. If there is an intermediate part other than the part, the thickness of the intermediate part is different.
The upper limit of the difference in thickness is not particularly limited because the effect can be exerted as it is larger. However, the formability of the stepped portion, the distance to the surface of the cell culture portion, etc. It can be set as appropriate according to the size of the cell culture container.

  The difference in thickness means that the stepped portion may be thicker or thinner than the adjacent opening side member.

  The shape of the stepped portion in the present invention is not particularly limited as long as the thickness is different at least at the boundary portion with the adjacent opening side member. Is preferably a fixed shape. This is because the step portion can be easily formed by the above shape.

  The material constituting the stepped portion in the present invention is not particularly limited as long as the stepped portion can be formed with a desired thickness, and the same material as the lid base material described later, or the above hydrophobic property Although it can be the same as the material which comprises a part and a water absorption part, it is preferable that it is the material which comprises a hydrophobic part especially, ie, a level | step difference part is also a hydrophobic part. This is because the movement of water can be more stably prevented.

  The method for forming a stepped portion in the present invention is not particularly limited as long as it can form a stepped portion having a desired thickness. For example, a method similar to that for the hydrophilic portion can be used.

(D) Water blocking unit The water blocking unit in the present invention is not particularly limited as long as it can stably prevent the movement of water, and may be one type or a combination of two or more types. It may be. For example, it may be composed of only the hydrophobic part, the water absorbing part or the step part as described above, or a combination thereof, more specifically, as shown in FIG. 10 or FIG. However, a hydrophobic part 21 is formed adjacent to the water film forming part 11 and has a water absorbing part 22 or a step part 23 so as to surround the hydrophobic part 21, or a hydrophobic part as shown in FIG. 21, a stepped portion 23, and a water absorbing portion 22.
10 to 12 indicate the same members as those in FIG. 1, and a description thereof will be omitted here.

(3) Lid part The lid part in this invention covers a cell culture part, and has the said water film formation part and water cutoff part in an inner surface. Further, at least during observation, the water film forming part and the water blocking part are formed so as to cover the opening of the cell culture part and the upper surface of the side wall, respectively.
In the present invention, when the lid portion covers the cell culture portion and can move with respect to the cell culture portion in a plan view, even if such movement occurs, It is preferable that the film forming portion and the water blocking portion are formed so as to overlap the predetermined region in plan view.

The lid portion in the present invention may be composed of the water film forming portion and the water blocking portion, and the lid base material and the water film forming portion and the water blocking portion formed on the inner surface of the lid base material. It may be.
Moreover, the cover part in this invention may have other structures other than a water film formation part and a water interruption | blocking part as needed.

In the present invention, the material constituting the lid substrate is not particularly limited as long as it has transparency and can stably form the water film forming portion and the water blocking portion. It can be the same as the material which comprises the cover part in a cell culture container. Specifically, it can be the same as the material constituting the cell culture section.
That is, a lid such as a petri dish or a multiwell plate that is commercially available as a general cell culture container can be used as the lid base material.

The shape of the lid in plan view in the present invention can be the same as that of a general cell culture container such as a circle or a polygon as shown in FIG.
Moreover, as a cross-sectional view shape of a cover part, as shown in FIG.1 and FIG.2 already demonstrated, it usually has a cover side part which covers a part of side surface of the said cell culture part.
In the present invention, when threading is performed on the outer peripheral portion of the cell culture portion, such inner surface of the lid side surface portion may be threaded.

3. Cell Culture Container The cell culture container of the present invention has at least the cell culture part and the lid part, and may have other structures as necessary.
As such other configurations, for example, as illustrated in FIG. 13, when the cell culture container of the present invention has a plurality of cell culture units, a dish unit 31 that supports the plurality of cell culture units is given. Can do.
In addition, about the code | symbol in FIG. 13, since it shows the same member as the thing in FIG. 1, description here is abbreviate | omitted.

  As cells cultured using the cell culture container of the present invention, various cells can be used. For example, it may be an anchorage-dependent cell that adheres to a scaffold such as a concave surface of a cell culture part. Suspension cells that do not have dependence may be used.

  The present invention is not limited to the above embodiment. The above-described embodiment is an exemplification, and the present invention has substantially the same configuration as the technical idea described in the claims of the present invention, and any device that exhibits the same function and effect is the present invention. It is included in the technical scope of the invention.

  Hereinafter, the present invention will be specifically described with reference to examples.

[Example]
A polystyrene sheet having a high anti-fogging function due to its high hydrophilicity (Sandick sheet 210, 300 μm thick) is cut out to 32 mmφ, and placed on the center of the lid of a 35 mmφ cell culture dish made by BD Affixed. PDMS (polydimethylsiloxane: Sylgard (registered trademark) 184 manufactured by Dow Corning) was used for pasting. PDMS before curing was applied to the petri dish lid surface, a polystyrene sheet was placed on the PDMS surface, and the mixture was allowed to stand overnight for pasting. The petri dish thus obtained was placed in an incubator at 37 ° C. for 1 hour, then taken out and allowed to stand at room temperature. As a result, a water film was formed on the polystyrene sheet, and a good observation region could be secured. On the other hand, since the petri dish cover material is exposed in the surrounding area where the polystyrene sheet is not affixed, a water film was not formed. In addition, since the attached polystyrene sheet was 300 μm thick, there was a 300 μm step between the lid surface and the polystyrene sheet, thereby confirming that the water film was limited within the range on the polystyrene sheet.
In addition, the water contact angle of each material was as follows.
・ Polystyrene sheet: 10 ° or less ・ Inner surface of cell culture dish: 80 °

DESCRIPTION OF SYMBOLS 1 ... Side wall 2 ... Recessed part 3 ... Cell culture part 10 ... Lid part 11 ... Water film formation part 12 ... Water blocking part 13 ... Lid base material 20 ... Cell culture vessel 21 ... Hydrophobic part 22 ... Water absorption part 23 ... Step part 31 ... Plate part

Claims (6)

A cell culture part having a recess surrounded by a side wall;
A lid that covers the cell culture part;
A cell culture vessel having
The inner surface of the lid portion is formed so as to surround the water film forming portion and the water film forming portion formed so as to overlap a part of the opening of the concave portion of the cell culture portion in plan view, and A cell culture vessel comprising a water blocking portion formed so as to overlap the upper surface of the side wall in plan view.   The cell culture container according to claim 1, wherein the water film forming part is a hydrophilic part.   The cell culture container according to claim 1, wherein the water film forming part is a fine uneven part.   The cell culture container according to any one of claims 1 to 3, wherein the water blocking part includes a hydrophobic part.   The cell culture container according to any one of claims 1 to 4, wherein the water blocking part includes a water absorbing part.   The cell culture container according to any one of claims 1 to 5, wherein the water blocking part includes a step part.

Images