JP2011169655A - Stain solution and staining method - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for rapidly performing eosin staining and finding out a stain solution reduced in irritant smell to reduce the use amount of the stain solution. <P>SOLUTION: An eosin stain solution is prepared by adding 5-25% of lactic acid and 0.5-10% of acetic acid to eosin dissolved in a mixed solution of water and a water-soluble organic solvent. One of a cylinder and a holder having a hole is closely fixed to a tape, to which a square layer obtained by a tape stripping method is adhered, and the eosin stain solution is put in the cylinder or the hole to perform staining. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to a human and environmentally friendly dyeing solution and method that can be carried out quickly and anywhere.

One way to know the skin quality is to stick the adhesive tape etc. to the skin surface, peel it off, transfer the skin stratum corneum cells to the adhesive surface such as the adhesive tape, and attach the skin corner attached to the adhesive surface such as the adhesive tape. If necessary, the layer cells are further copied onto a glass slide and stained.
A staining method used for this is eosin staining.

Eosin is an acidic dye, and since the dye itself is negatively charged, it binds and dyes positively charged cytoplasm, cytoplasm, and fibers. The staining property changes due to the difference in pH, and the positive charge increases due to the change on the tissue side under acidic conditions, and it becomes easier to bind to the negatively charged dye. For this reason, use and dyeing time can be shortened in the state where acetic acid was added and dyeability increased.
Such dyeing is a work that is very often carried out nowadays when a lot of cosmetics corresponding to each person's skin quality are known in detail, and the dyeing process is equipped as before There is a need not only to take a certain amount of time in a laboratory or the like, but also to quickly dye at a store selling cosmetics and select a product based on the result. In addition, the environment is not well ventilated, and an unspecified number of people come into contact.

If the amount of acetic acid is increased in order to shorten the dyeing time as described above, the odor of acetic acid becomes stronger, the equipment is not well-equipped, and it is an odor that cannot be tolerated by general consumers in stores that sell cosmetics.
That is, the present inventors are to find a staining solution that has a low acetic acid odor and can be quickly dyed, and a staining method using this staining solution.
Furthermore, dyeing is usually performed using a dyeing vat with a large amount of dyeing solution, dyeing, enlarging, and observing with a microscope, etc. The purpose is to find an environmentally friendly dyeing method.

As a result of intensive studies, the present inventors have found that the following method solves this problem.
Dissolve eosin in the required amount of solvent. As the solvent, water, a water-soluble organic solvent such as ethanol, or a mixture thereof is used.
However, ethanol: water = 50: 50 to 99: 1 is usually used.
On the other hand, 5 to 25% by volume, preferably 8 to 20% lactic acid is added and stirred.
Further, acetic acid is added in an amount of 0.5 to 10% based on the amount of eosin dissolved in a mixture of the first water and the water-soluble organic solvent.
As a result of investigations by the inventors, it was possible to find a liquid having an acceptable acetic acid odor and a good dyeing state by adding a small amount of acetic acid because the dyeing state was poor with lactic acid alone.
A method of using this staining solution quickly and using a small amount of staining solution was considered.
In this method, one of a prepared cylinder or a holder having a hole was tightly fixed to this tape by using the adhesive force of the tape to which the stratum corneum cells obtained by the tape stripping method were applied.

A cylindrical holder or a holder having a hole will be described below.
The cylinder may be a cylinder or an oval cylinder, but a cross-sectional shape may be a triangle, quadrangle, or other polygonal cylinder, but a cylinder having a cross section that is not wider than the width of the tape is better. After that, there is little leakage of the dyeing solution to be put into the cylinder, and it is meaningless to dye the part without the tape. That is, the size of the cross section is 10 to 1000 mm ² , preferably 20 to 500 mm ² , and if the cross section is small, the area where the stratum corneum is dyed is small, making it difficult to diagnose the target skin quality, If extra staining solution is required and the accuracy of the skin quality diagnosis is within a certain area, it will hardly help improve accuracy beyond that.
Further, there may be a flange on one or both cross sections of the tube.
The height (length) of the tube is preferably about 3 to 20 mm, and if it is too short, a necessary amount of the staining solution cannot be put into the tube and the staining solution may spill.

Alternatively, it is not a cylinder, but a holder having a hole may be used. It should be understood that there is a hole that can be put put dyeing solution in a plate with a certain thickness, since the cross section of the hole is an area that can be dyed, as with the tube, 10 to 1000 mm ^2, preferably 20 to 500 mm ² Any shape can be selected as long as the shape of the hole is within the range in which the staining status can be confirmed.
Since the thickness of the holder has the same meaning as the height (length) of the cylinder, about 3 to 20 mm is preferable.
The shape of the holder is not particularly limited as long as the shape of the holder is plate-like and the tape to which the horny layer cells are attached can be closely fixed.
In addition, the hole does not need to penetrate both sides of the holder, and may be a recess. In this case, before the tape with the horny layer cells adhered is firmly fixed to the recess, the staining solution is put in the dent, After the attached tape is closely fixed to the recess, it is inverted and the stratum corneum cells are brought into contact with the staining solution.
There is no problem as long as the material of the holder having the tube and the hole is a material that does not crack or dissolve by the staining solution.

Such a cylinder is closely fixed to the tape. This is because the staining solution is put into the tube, and the tape and the tube need to be closely fixed so as to secure the necessary time and the time for the staining solution to contact the stratum corneum cells attached to the tape. In consideration of recovery of the staining solution, it is better that the staining solution is closely fixed so that the staining solution does not leak.
For this reason, if a holder with a tube or hole, a tape with stratum corneum cells attached, a plate-like elastic body, and a plate-like body with little deformation are installed in this order, the stain solution will be less likely to leak if they are brought into close contact with some pressure. .

The time for which the staining solution contacts the stratum corneum attached to the tape varies depending on the staining solution, but is 30 seconds to 15 minutes, preferably 2 to 10 minutes, from the gist of the present invention. If it is short, the dyeing may be insufficient, and if it is long, the dyeing can be done in a short time, which is the gist of this patent.
Needless to say, the working time of the dyeing solution varies depending on the acid concentration of the dyeing solution.
After the staining time has elapsed, the staining solution is removed, and observation is performed using means such as a microscope or a microscope. Also, these images are obtained and necessary analysis is performed.

Comparative test A.1. Staining solution The staining solution prepared as shown in Table 1 was used.

B. Execution method The tape which the horny layer cell adhered was obtained using the cellophane tape (Nichiban company make, brand name cellophane adhesive tape CT405AP-18) from a human cheek part.
The cross-section side with the flange of the cylinder of FIG. 2 was pressed against the adhesive surface of this tape and set on the rubber plate of the dyeing tool of FIG. The pressing plate of the dyeing tool was pressed and fixed with a stopper so as to press the flange of the cylinder.
200 μL of the staining solution prepared as described above was placed in this tube and allowed to stand for 5 minutes.
Thereafter, the dyeing solution was discarded, lightly washed with water, and the dyed tape was adhered to the slide glass.

As a result, as shown in FIGS. 5 to 8, when lactic acid and acetic acid are used in a specific ratio in the staining solution, the staining is good and can be dyed in a short time, and the odor of acetic acid is greatly reduced compared with the case where only acetic acid is used. did it. Furthermore, by using the dyeing tool of FIGS.

Is the dyeing tool used in the comparison test. Stopper 2.20 × 20mm 2mm thick rubber plate (set the tape attached on it) 3. Hinge 4. 4. Tube installation position Holding plate Is a tube that contains the staining solution used in the comparative test (photo taken from the opposite side of the set tape with stratum corneum cells attached) Is a tube for storing the staining solution used in the comparison test (the tube is seen from the side with a flange on one side, and the flange and the tape on which the stratum corneum cells are adhered are firmly fixed by the pressing plate of 5 in FIG. ) When the tape with stratum corneum cells attached is set with the staining tool used in the comparative test and fixed tightly Image stained with staining solution of Example 1 in comparative test Image stained with staining solution of Example 2 in comparative test Image stained with the staining solution of Comparative Example 1 in the comparative test Image stained with the staining solution of Comparative Example 2 in the comparative test

Claims (3)

  An eosin staining solution obtained by adding 5 to 25% lactic acid and further adding 0.5 to 10% acetic acid to eosin dissolved in a mixture of water and a water-soluble organic solvent.   A stratum corneum staining method using the eosin staining solution of claim 1   The dyeing method according to claim 2, wherein one of the holder having a cylinder or a hole is closely fixed to the tape with the stratum corneum obtained by the tape stripping method, and the eosin staining solution of claim 1 is put into the cylinder or the hole.