JPS63113358A - Keratin layer inspection - Google Patents

Abstract

PURPOSE:To obtain a clear and well-preservable specimen, by a method wherein a keratin layer is adhered to a sticky substance applied on a tape, this tape is stuck on a transparent plate previously coated with a fastening agent and then, is removed after the plate is immersed into a solvent. CONSTITUTION:First, a tape previously coated with a sticky substance is put tight on human skin and then, peeled off to sample a part of a keratin layer on the tape. Then, the tape is stuck on a transparent plate previously coated with a fastening agent. Thereafter, the transparent plate with the tape stuck thereon is immersed for 1-3hr into a solvent adapted to solve the sticky substance applied on the tape to enhance the peeling performance of the tape and then, the tape is peeled off the transparent plate carefully. Then, by carrying out a balsam sealing or other operations, a keratin layer specimen is made and the specimen thus obtained is observed with an optical microscope or the like. This provides a clear and well-preservable specimen thereby enabling the inspection of a keratin layer accurately and easily.

Description

[Detailed Description of the Invention] [Field of Industrial Application] The present invention relates to a method for inspecting the stratum corneum of the skin, and can be used in fields such as cosmetics, foods, medicines, and skin quality inspection. It is.

[Conventional technology]

Essentially, when selling products such as cosmetics that are required to maintain good skin condition, it is necessary to accurately understand the user's skin condition and provide guidance on how to apply makeup accordingly. Based on this idea, in recent years, it has become an important issue to know the skin quality, skin texture, skin condition, etc. by observing or measuring the skin condition.

Conventionally, many methods have been proposed to observe these skin conditions, but it has been found that accurate skin examinations can be performed by closely observing the stratum corneum on the skin surface. Conventionally, methods for examining the surface condition of the skin include (a) a method of directly observing the skin with the naked eye;

(b) Sump method (a method in which an organic solvent such as amyl acetate is dropped onto a celluloid plate, the celluloid plate is brought into close contact with the skin, and then it is peeled off to create a skin replica and observed).

(C) A method in which an enlarged image of the skin is displayed on a monitor and observed using a device such as a videoscope.

Such methods have been used.

[Problem that the invention seeks to solve]

However, method (a) or (C) is based on external appearance, and although it is convenient for measuring skin color and luster,
It was not possible to precisely examine the condition of the skin. Also,
Although the method (b) is an advantageous method for examining the roughness and unevenness of the skin, it merely measures the surface shape. Therefore, in order to examine the condition of the skin in more detail, the best method is to collect the stratum corneum and observe it.

This stratum corneum inspection method includes, for example, applying a known adhesive to a transparent plate, bringing it into close contact with the skin, and then peeling it off to retain the stratum corneum on the transparent plate and using it as a sample of the stratum corneum. Conceivable.

However, with this method, it is not possible to obtain a clear and well-preserved stratum corneum specimen, and the test subject's skin is often damaged.

Therefore, it is difficult to precisely observe or inspect the condition of the skin.

[Means for solving problems]

The present invention was developed in view of the above-mentioned problems, and includes a tape coated with an adhesive substance soluble in an organic solvent and the stratum corneum of the skin adhered to the adhesive substance, which is coated with an adhesive in advance. We focused on the fact that a clear and long-lasting specimen of the stratum corneum can be obtained by attaching it to a transparent plate, then peeling off the adhesive tape using an organic solvent, and fixing the stratum corneum to the transparent plate. By observing and measuring the stratum corneum cells using this stratum corneum specimen, the condition of the skin can be investigated precisely and quickly, and the above problems can be solved.

[Effect]

In the method of the present invention, by taking the above-described measures, the stratum corneum can be collected from the skin using a tape coated with an adhesive substance soluble in an organic solvent.

The stratum corneum specimen applied to the present invention is a type of stratum corneum specimen that is made by fixing the stratum corneum, which is a thin film-like cell group that exists in the outermost layer of the epidermis that covers the surface of human skin, on a transparent plate. This is a tissue specimen. The above-mentioned stratum corneum specimen allows the stratum corneum cells to be observed in detail by staining, etc., and is particularly suitable for microscopic observation. If the stained stratum corneum cells have nuclei, they can be clearly observed;
Since the keratinization state of epidermal cells can be determined, it is also possible to know the metabolic rate of epidermal cells and the state of rough skin. Furthermore, by measuring the area, circumference, aspect ratio, homogeneity, etc. of the stratum corneum cells, the degree of aging and skin quality of the skin can be objectively evaluated. Of course, the above-mentioned stratum corneum specimen can be enlarged on a screen using equipment such as a videoscope.
It is also suitable to observe and measure this.

In this way, a lot of knowledge about the skin condition can be obtained by using the method of the present invention, and since it is possible to accurately understand the skin condition of each individual, based on this, it is possible to provide makeup guidance according to the person's skin type. It is also possible to do so.

Next, the adhesive substance used to prepare the stratum corneum specimen according to the present invention is a resin or polymer compound that is soluble in an organic solvent, and specific examples include polyvinyl acetate, polyvinyl acetate,
Polyvinyl alcohol, polyvinyl acecool, polyvinyl chloride, polyvinylidene chloride, polyvinyl ether,
Polybutene, asphalt, natural rubber or its derivatives, SBR rubber, NBR rubber, butadiene-vinylpyridine rubber, butyl rubber, chloroprene rubber,
Recycled rubber, cyanoacrylate rubber, gum arabic,
Rubber materials such as tragacanth rubber are listed, and these 1
A species or a mixture of two or more species may be used alone or in combination with another base material. Among the above-mentioned adhesive substances, natural rubber or its derivatives, SBR-based com, N
Rubber materials such as BR rubber, butadiene-vinylpyridine rubber, butyl rubber, chlorobrene rubber, recycled rubber, cyanoacrylate rubber, gum arabic, rubber tragacanth, etc. have good removability when removing the tape and do not change the stratum corneum cells. are preferably used in the method of the present invention. Of course, in the method of the present invention, tapes coated with the above-mentioned adhesive substances, such as commercially available cellophane tapes, are also suitably used. However, tapes such as duct tape that use gummy substances as adhesive substances cannot be used in the method of the present invention because the gummy substances are insoluble in the above-mentioned organic solvents. It should be noted that it is sufficient to use the material of the tape itself that is commonly or industrially used, and in addition to those with a uniform thickness, those with pores, those with a mesh shape, etc. can also be used. .

Further, the organic solvent used when preparing the stratum corneum specimen according to the method of the present invention is one that can dissolve the above-mentioned sticky substance, and specific examples of such organic solvents include methanol, Ethanol, propatool, butanol, ether, acetone, chloroform, hexane, T
HF, ethyl acetate, 0-xylene, m-xylene, p-
Examples include xylene, methylbenzene, ethylbenzene, propylbenzene, benzene, toluene, amyl acetate, etc., and one or more of these may be used as a mixture or in combination.

Next, the fixing agent used when preparing the stratum corneum specimen according to the method of the present invention is one that fixes the stratum corneum to the transparent plate, does not degenerate the stratum corneum cells, and does not contain the above-mentioned organic It is not easily soluble in solvents. Examples of such a fixing agent include known adhesives, proteins, etc., but known adhesives are preferably used because the obtained stratum corneum specimen is clearer, and specific examples include: Polyvinyl adhesives such as polyvinyl acetate, polyvinyl alcohol, polyvinyl acetal, polyvinyl chloride, polyvinylidene chloride, polyvinyl ether, melamine resin adhesives, phenolic resin adhesives, resorcinol adhesives, xylene resin adhesives, furan resin adhesives adhesive, polyisocyanate resin adhesive, polyester resin adhesive, polyamide adhesive, polybutene adhesive, ethylene copolymer adhesive, thermoplastic polyester adhesive, polyester acrylate adhesive, silicone rubber adhesive, phenol mixed adhesive adhesives, epoxy adhesives, cyanoacrylate adhesives, anaerobic polyester adhesives, polybenzimidazole adhesives, polyimide adhesives, polyimide amide adhesives, Trenise adhesives, metalloxane adhesives, glue adhesives agent, casein adhesive, cellulose adhesive, viscose adhesive, etc., and these adhesives may be used alone or in combination of two or more. Among these adhesives, polyvinyl adhesives are most preferred because they are resistant to staining when dyed. Examples of the above proteins include ovalbumin, milk albumin, myogen, serum albumin, globin, albumins such as leucosin, serum globulin, β-lactoglobulin, lysozyme, myosin, edestin, insulin, fibrinogen, thyroglobulin, etc. globulins, soluble collagen, elastin, keratin,
Silk fibroin, spondin, butadiene, histone, protamine, egg white, gelatin, peptone, peptide,
Other examples include phosphoproteins, lipoproteins, glycoproteins, etc., and one or a mixture of two or more of these may be used alone or in another base material such as a polyhydric alcohol.

Further, the transparent plate used when preparing the stratum corneum specimen according to the method of the present invention is transparent and does not interfere with observation of the stratum corneum specimen, and is insoluble or sparingly soluble in organic solvents. Specific examples include glass plates and resins such as polymethylpentene and polyethylene, and among these, glass plates are most preferably used in terms of transparency and solvent resistance.

In the method of the present invention, first, a tape coated with the above-mentioned adhesive substance is brought into close contact with the skin, and then a part of the stratum corneum can be collected on the tape by peeling it off. Next, this tape is attached to a transparent plate that has been previously coated with the above-mentioned adhesive. The important thing here is to apply a fixing agent to the transparent plate in advance. If this pretreatment is not done, the stratum corneum will be released from the transparent plate during the steps such as peeling off the tape described below. It is inappropriate because it will cause Also, when attaching a tape holding the stratum corneum to a transparent plate, the width of the tape should be 3 to 10 u.
It is preferable that the degree of If the tape is too wide, it has poor permeability to organic solvents and is inconvenient to handle. Next, the transparent plate to which the above tape was pasted was immersed in the above-mentioned organic solvent for 1 to 3 hours to dissolve or swell a portion of the adhesive substance adhering to the tape, thereby increasing the releasability of the tape. After that, carefully peel it off from the transparent plate. Furthermore, it is also suitable to continue immersion in an organic solvent for the purpose of removing residual sticky substances.

After the stratum corneum is transferred to a transparent plate, it is usually dyed and dyed using conventional methods.
Perform operations such as balsam encapsulation, use this as a stratum corneum specimen, and examine the stratum corneum using methods such as (a) and (b) below to use as material for skin quality testing.

(b) Observe the obtained stratum corneum specimen with an optical microscope,
Examine the condition, texture, and quality of the skin based on the shape, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells.

(b) The obtained stratum corneum specimen is displayed on a monitor using a videoscope, and the skin condition and skin properties are determined based on the shape, quantity, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells. ,
The skin condition, skin quality, skin quality, degree of skin aging, age of the skin, etc. are investigated by examining the skin quality, and further measuring the area, circumference, aspect ratio, etc. of the stratum corneum cells.

〔Example〕

Next, examples of the method of the present invention will be shown.

<Example-1> A piece of cellophane tape (cut from commercially available adhesive cellophane tape) with a width of 121 m and a length of 20 degrees is pasted on the cheek, held for a while, and then gently peeled off and cut with scissors. It was made to have a width of approximately 6 cm and a length of 20 zm, and was attached to a glass slide that had been uniformly coated with a polyvinyl acetate adhesive.

Next, this glass slide is immersed in xylene for 2 hours. Next, after gently peeling off the tape using tweezers, the sample was further immersed in xylene for 1 hour.

Then, after taking it out and drying the xylene, gentian violet-brilliant green staining solution (gentian violet 19 and brilliant green 0) was used.
．． 59 dissolved in distilled water 1009)
After soaking for 0 minutes, washing with water, drying, and enclosing in balsam according to the usual method, a stratum corneum specimen is obtained.

Next, the obtained stratum corneum specimen was observed using an optical microscope,
Skin condition, skin texture, skin quality, etc. were investigated based on the shape, quantity, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells.

The obtained stratum corneum specimen is an unprecedentedly clear specimen of the stratum corneum, and as shown in Figure 1, the stratum corneum cells can be clearly observed, and in particular, the normal or abnormal state of keratinization can be observed in detail. Therefore, we were able to investigate skin metabolism, aging degree, skin condition, etc.

<Example 2> A piece of cellophane tape (cut commercially available adhesive cellophane tape is used) with a width of 12 x m and a length of 20 x is pasted on the cheek, pressed for a while, and then gently peeled off and cut with scissors. It has a width of approximately 6 cm and a length of 20 rtutr, and is attached to a glass slide that has been uniformly coated with polyvinyl acetate adhesive in advance.

Next, the slide glass is immersed in xylene for 2 hours, the tape is gently removed using tweezers, and then the slide glass is further immersed in xylene for 1 hour. Next, take it out and dry the xylene, then add gentian violet-brilliant green staining solution (gentian violet 19 and brilliant green 0.59 to 10 parts distilled water.
After 5 to 10 minutes of washing and drying, the specimen is encapsulated in balsam by a conventional method to obtain a stratum corneum specimen.

Next, the obtained stratum corneum sample is displayed on a monitor using a videoscope, and the skin condition and skin quality are determined based on the shape, quantity, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells. ,
By examining the skin quality, etc., and measuring the area, circumference, aspect ratio, etc. of the stratum corneum cells using an image analysis device, we can determine the skin condition, skin type, skin type, degree of skin aging, and skin age. etc. were investigated.

In the obtained stratum corneum specimen, stratum corneum cells were clearly observed as in Example 1, and skin metabolism, degree of aging, skin condition, etc. could be investigated.

<Example 3> A piece of cellophane tape with a width of 12 mm and a length of 20 mm (commercially available adhesive cellophane tape is cut and used) is pasted on the cheek, pressed for a while, and then gently peeled off and cut with scissors. The width was about 6 cm and the length was 20 x m, and it was carefully pasted onto a glass slide that had been uniformly coated with a mixture of ovalbumin:glycerin = l:l (weight ratio).
Leave at a temperature of 60°C for 2 hours.

Next, this glass slide was immersed in ethanol for 10 minutes, dried, and further immersed in xylene for 2 hours.
Gently remove the tape using tweezers and then soak in xylene for an additional hour. Next, take it out and dry the xylene, then gentian violet-brilliant green staining solution (gentian violet 1
9 and 0.5g of Brilliant Green in 100g of distilled water
After 5 to 10 minutes of washing and drying, the specimen is encapsulated in balsam by a conventional method to obtain a stratum corneum specimen.

Next, the obtained stratum corneum sample is displayed on a monitor using a videoscope, and the skin condition and skin quality are determined based on the shape, quantity, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells. ,
By examining the skin quality, etc., and measuring the area, circumference, aspect ratio, etc. of the stratum corneum cells using an image analysis device, we can determine the skin condition, skin type, skin type, degree of skin aging, and skin age. etc. were investigated.

In the obtained stratum corneum specimen, stratum corneum cells were clearly observed as in Example 1, and skin metabolism, degree of aging, skin condition, etc. could be investigated.

<Example-4> A paper tape with a width of 10xx and a length of 20'1111, evenly coated with an adhesive substance containing gum arabic, gum tragacanth, etc., is pasted on the cheek, held for a while, and then gently rubbed. Peel it off and use scissors to make it about 6cm wide.
, 20 mm in length, and attached to a glass slide to which a cellulose adhesive was evenly applied in advance.

Next, the slide glass is immersed in acetone for 2 hours, the tape is gently removed using tweezers, and then the slide glass is further immersed in xylene for 1 hour. Next, it is taken out and dried with xylene, and then stained with hematoxylin and eosin. After drying, it is sealed in balsam by a conventional method to obtain a stratum corneum specimen. Next, the obtained stratum corneum specimen is observed using an optical microscope, and the shape, quantity, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells are examined to determine the skin condition, skin texture, and skin quality. etc. were investigated.

In the obtained stratum corneum specimen, stratum corneum cells were clearly observed as in Example 1, and skin metabolism, degree of aging, skin condition, etc. could be investigated.

<Example-5> A plastic tape with a width of lO■ and a length of 20 ryua, evenly coated with an adhesive substance containing natural rubber, was pasted on the outer part of the upper arm, and gently peeled off.
Thereafter, it is treated in the same manner as in Example 1 and stained with Giemsa to obtain a stratum corneum specimen. Next, the obtained stratum corneum specimen is displayed on a monitor using a videoscope, and the shape and quantity of the stratum corneum cells are determined.
Examine the skin condition, texture, skin quality, etc. from size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. Furthermore, use an image analysis device to examine the area, circumference, oblateness, etc. of the stratum corneum cells. By measuring the skin condition, skin quality, skin quality, degree of skin aging, skin age, etc., were investigated.

In the obtained stratum corneum specimen, stratum corneum cells were clearly observed as in Example 1, and skin metabolism, degree of aging, skin condition, etc. could be investigated.

<Example-6> Width 1011 m, length 20 r with uniformly distributed pores
NX cellophane tape (processed commercially available adhesive cellophane tape is used) is pasted on the inner side of the upper arm, gently peeled off, and treated in the same manner as in Example 2 to obtain a stratum corneum specimen. Next, the obtained stratum corneum sample is displayed on a monitor using a videoscope, and the skin condition and skin quality are determined based on the shape, quantity, size, uniformity, homogeneity, degree of aggregation, presence or absence of nuclei, etc. of the stratum corneum cells. By examining the skin quality, etc., and measuring the area, circumference, flattening ratio, etc. of the stratum corneum cells using an image analysis device, we can determine the skin condition, skin type, skin type, degree of skin aging, and skin condition. We looked into age, etc.

In the obtained stratum corneum specimen, the stratum corneum cells were clearly observed as in Example-①, and skin metabolism, degree of aging, skin condition, etc. could be investigated.

〔Effect of the invention〕

By having the above-described structure, the present invention tests the stratum corneum using a specimen that is clear, easy to observe, and has a good shelf life. Therefore, the test of the stratum corneum can be performed reliably and easily, and the skin quality etc. It can be effectively used for inspection.

[Brief explanation of the drawing]

Figure 1 shows a stratum corneum specimen applied to the method of the present invention (Example-
1) is an optical micrograph (magnification approximately 700 times).

Claims (5)

[Claims] (1) A tape coated with an adhesive substance soluble in an organic solvent and with the stratum corneum of the skin attached to this adhesive substance is attached to a transparent plate coated with an adhesive in advance, and then After soaking in an organic solvent, a part of the stratum corneum is substantially fixed on a transparent plate by removing the tape, and if necessary, this is stained to examine the condition of the stratum corneum using the prepared stratum corneum specimen. A stratum corneum inspection method characterized by: (2) The adhesive substance is a rubber material such as natural rubber or its derivatives, SBR rubber, NBR rubber, butadiene-vinylpyridine rubber, butyl rubber, chloroprene rubber, recycled rubber, cyanoacrylate rubber, gum arabic, rubber tragacanth, etc. The method for inspecting the stratum corneum according to claim 1, wherein the method is one or a mixture of two or more selected from the following. (3) The stratum corneum inspection method according to claim 1, wherein the tape coated with an adhesive substance soluble in an organic solvent is cellophane tape. (4) The fixing agent is one or a mixture of two or more selected from polyvinyl adhesives such as polyvinyl acetate, polyvinyl alcohol, polyvinyl acetal, polyvinyl chloride, polyvinylidene chloride, and polyvinyl ether. The method for inspecting the stratum corneum according to claim 1, characterized in that it is contained in a base material. (5) The stratum corneum inspection method according to any one of claims 1 to 4, wherein the transparent plate is a glass plate.