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Hydrogel thin film containing extracellular matrix components

a technology of extracellular matrix and hydrogel, which is applied in the field of hydrogel thin film containing extracellular matrix components, can solve the problems of difficult handling of collagen hydrogel itself, difficult preparation of cell culture substratum, and more simple utilization methods, so as to achieve convenient preparation for culturing, appropriate elastic strength, and easy handling

Inactive Publication Date: 2002-07-04
TAKEZAWA TOSHIAKI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0023] As described above, the hydrogel thin film containing an extracellular matrix components of the present invention, as the thin film itself and as that integrated with the retainer, has an appropriate elastic strength and a thin film shape permitting easy handling, and enables achievement of a biochemical composition as a cell culture substratum, thus providing easy preparation for culturing and an excellent expediency.
[0029] More particularly, in the manufacturing method of the present invention, the vitrified gel containing an extracellular matrix components such as collagen is hydrated. This provides a hydrogel thin film containing an extracellular matrix components having a satisfactory elastic strength, which serves as a cell culture substratum and is useful for preventing organ adhesion.
[0030] For vitrification, in the present invention, it is the general practice to slowly and completely dry the gel containing the extracellular matrix components having a terminal concentration in an aseptic manner (for example, through aseptic air drying), thereby vitrifying the gel. Hydration of the vitrified gel can easily be effected by treating it by PBS, for example.

Problems solved by technology

Regarding the culturing methods using collagen or the like attracting the general attention as to usage thereof, however, in the case of collagen hydrogel for example, there is a problem of difficulty in handling the collagen hydrogel itself because of the softness.
It is not therefore easy to prepare a cell culture substratum, and a more simple method for utilization has not as yet been established.

Method used

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  • Hydrogel thin film containing extracellular matrix components
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  • Hydrogel thin film containing extracellular matrix components

Examples

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Effect test

example 1

Preparation of Collagen Hydrogel Thin Film

[0032] 2 ml of quintuple-concentration Dulbecco's Modified Eagle Medium (GIBCO #31600-034), 0.1 ml of 10,000 units / ml penicillin and 10,000 .mu.g / ml streptomycin (GIBCO #15140-031), 0.2 ml of 1M HEPES (GIBCO #15630-023), 0.493 ml of 7.5% sodium bicarbonate solution (GIBCO #25080-011), 1.407 ml of distilled water, and 1 ml of fetal bovine serum were Put in a sterilized conical tube (Falcon #2070) chilled on ice, having an inner volume of 50 ml, and mixed. Then 4.8 ml of 0.5% aqueous type-I collagen solution (CELLGEN I-AC or I-PC, made by Koken Company) was added into the tube and uniformly mixed. After placing 2 ml of the mixed collagen solution having a final concentration of 0.24% in a culture dish made of hydrophobic Polystyrene (.O slashed.35 mm: Falcon #1008), the solution was held in a humidified incubator at 37.degree. C. in the presence of 5% CO.sub.2 / 95% air in 3 hours for its gelation. This collagen gel of the final concentration of...

example 2

Preparation of Collagen Hydrogel Thin-film with Peripheral Wire Retainer

[0033] A circular retainer (1) with a knob as shown in FIG. 2 was made with a stainless steel wire (size: #20; 0.9 mm), and 2 ml of the mixed collagen solution having a final concentration of 0.24% prepared in Example 1 was placed, together with this wire retainer, into a hydrophobic polystyrene culture dish (.O slashed.35 mm; Falcon #1008). In the same manner as in example 1, the collagen solution with the retainer was vitrified after its glation. The vitrified collagen gel was hydrated by adding 2 ml PBS. Further, the collagen gel thus hydrated was rinsed several times with 2 ml of PBS. The hydrated collagen gel could be peeled off and collected from the dish by slowly lifting the stainless steel knob, in the form of a thin-film collagen hydrogel having a satisfactory elastic strength with a wire retainer surround it as shown in FIG. 3.

example 3

Preparation of Gauze Embedding Type Collagen Hydrogel Thin Film

[0034] A sterilized type-III gauze as prescribed in the Japanese Pharmacopoeia (K-Pine made by, Kawamoto Hotai Zairyou Company) was cut in a circular shape with a knob in an aseptic manner, and immersed the cut gauze in a cell culture medium (Dulbecco's Modified Eagle Medium containing 10% fetal bovine serum, 20 mM HEPES, 100 units / ml penicillin and 100 .mu.g / ml streptomycin). The gauze was put, together with 5 ml of the mixed collagen solution having a final concentration of 0.24% prepared in Example 1, in a hydrophobic polystyrene culture dish (.O slashed. 60 mm; Falcon #1007). In the same manner as in example 1, the collagen solution with the gauze was vitrified after its gelation.

[0035] Furthermore, the vitrified collagen gel was hydrated by adding PBS in the same manner as in Example 1. The hydrated collagen gel could be peeled off and collected from the dish by slowly lifting the knob of the gauze, in the form of a...

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Abstract

The thin film of the invention comprises a hydrate of a vitrified gel containing an extracellular matrix components, which can be integrated with a retainer as required. A hydrogel thin film containing an extracellular matrix components such as thin-film collagen hydrogel thin film, which is useful for a cell culture substratum and for preventing organ adhesion, can be easily prepared, and is excellent in expediency.

Description

[0001] The present invention relates to a hydrogel thin film containing an extracellular matrix components. More particularly, the present invention relates to a novel hydrogel thin film containing an extracellular matrix components, which is useful for a cell culture substratum and prevention of organ adhesion, has an appropriate elastic strength, can be easily prepared and can be used simply.PRIOR ART AND PROBLEMS[0002] Cell culture has conventionally been carried out in various manners for various purposes including development of various medical techniques and various medical drugs.[0003] A method using an extracellular matrix components such as collagen is known as a method for cell culture. In this method, in the case of collagen for example, various trial efforts have been made to ensure a three-dimensionality as closest as possible to forms of biological tissues or functional expression by alleviating restrictions imposed as a two-dimensional planar culture for ordinary cell...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61L31/00A61L31/04A61L31/14C12N5/00C12N5/02
CPCA61L31/005A61L31/044A61L31/145C12N5/0068C12N2533/54
Inventor TAKEZAWA, TOSHIAKIYOSHIZATO, KATSUTOSHI
Owner TAKEZAWA TOSHIAKI
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