阅读说明：本技术 一种微小昆虫永久显微玻片标本制作方法 (Method for manufacturing permanent microscopic slide specimen of tiny insect ) 是由 党利红 李彦巧 王夏 于 2021-02-09 设计创作，主要内容包括：本发明公开了一种微小昆虫永久显微玻片标本制作方法,包括固定、脱色、脱水、固色、干燥和封片。本发明的有益效果在于,首先,发明的玻片标本制作方法相比于现有的标本制作方法步骤简单,使用材料不会造成环境污染且染色效果好,能够降低标本制作过程的复杂程度和对人体的伤害性；其次,本发明的玻片标本制作方法通过恒温真空室实现标本的干燥,易于冲体内溶液的充分蒸发,不会造成现有自然风干而出现的内部气泡,以及进一步造成的标本表面浑浊的问题；最后,本发明的玻片标本制作方法通过酸性品红酒精溶液对虫体染色,能够使虫体的身体结构更加明显的显现,通过不同浓度酒精的逐层脱水,不会引起虫体结构的收缩和变形。(The invention discloses a method for manufacturing a permanent microscopic slide specimen of a tiny insect, which comprises the steps of fixing, decoloring, dehydrating, fixing color, drying and mounting. The method has the advantages that firstly, compared with the existing specimen preparation method, the method for preparing the slide specimen has simple steps, does not cause environmental pollution by using materials, has good dyeing effect, and can reduce the complexity of the specimen preparation process and the harm to human bodies; secondly, the method for manufacturing the slide specimen realizes the drying of the specimen through the constant-temperature vacuum chamber, is easy to fully evaporate the solution in the rinsing body, and can not cause the problems of internal bubbles caused by the existing natural air drying and further turbidity on the surface of the specimen; finally, the method for manufacturing the slide specimen can enable the body structure of the worm to be more obviously displayed by dyeing the worm with the acid fuchsin alcohol solution, and the shrinkage and deformation of the body structure can not be caused by the layer-by-layer dehydration of the alcohol with different concentrations.)

1. A method for manufacturing a permanent microscopic slide specimen of a tiny insect is characterized by comprising the following steps:

step 1, soaking living insects in 65-76% alcohol solution;

step 2, taking out the insects from the alcohol solution, cutting the insects into bodies, putting the insects into a beaker added with a sodium hydroxide solution, putting the beaker on a heating plate, heating and soaking the insects, and cleaning the insects and the internal organs until the insects become transparent;

step 3, taking the polypide treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the polypide into constant-temperature cold bath acid fuchsin alcohol solution with gradually increased concentration for color fixation;

step 4, moving the polypide processed in the step 3 out, placing the polypide on filter paper, and placing the polypide in a constant-temperature vacuum chamber for drying and dehydration;

step 5, spraying xylene vapor to the inner and outer surfaces of the insects in the vacuum chamber in the step 4 to form a coating of 5-10 microns;

and 6, dripping Canadian gum on the glass slide, placing the insects treated in the step 5 on the Canadian gum, laying all parts of the insect body, covering a cover glass, and airing to obtain a permanent glass slide specimen.

2. The method as claimed in claim 1, wherein the concentration of the sodium hydroxide solution is 5%.

3. The method for preparing a permanent microscopic slide specimen for tiny insects as claimed in claim 1, wherein in step 2, the heating temperature of said heating plate is 100-120 ℃ and the heating time is 20 min.

4. The method for preparing a permanent microscopic slide specimen for tiny insects as claimed in claim 1, wherein in step 3, said acid fuchsin alcohol solution uses 30%, 55%, 65%, 90%, 100% alcohol as solvent.

5. The method for preparing a permanent microscopic slide specimen of tiny insects as claimed in claim 4, wherein in step 3, the insects are immersed in said acid fuchsin alcohol solution with alcohol solvent concentration of 30%, 55%, 65%, 90% for 40-80 min, respectively, and immersed at 100% for 80-160 min.

6. The method for manufacturing the permanent microscopic slide specimen for the tiny insects as claimed in claim 1, wherein the color fixing time of step 3 is 4-8 h.

7. The method for preparing a permanent microscopic slide specimen of tiny insects as claimed in claim 1, wherein the temperature of the constant temperature vacuum chamber in step 4 is 25 ℃, and the insects are dried and dehydrated on the filter paper until there is no solvent on the filter paper.

Technical Field

The invention relates to the technical field of insect specimens, in particular to a method for manufacturing a permanent microscopic slide specimen of a tiny insect.

Background

When morphological research of small insects (such as scale insects, aphids, whiteflies, mites and the like) is carried out in natural science research, technical popularization research, biological experiment teaching and extracurricular scientific and technological activities, in order to better understand the morphological structure of the small insects, the small insects are generally made into microscope slide specimens, and the internal structure of the insects is directly observed through a microscope. The microscope slide includes: both temporary and semi-permanent slides, permanent slides.

At present, common methods for manufacturing permanent microscope slides include a Venice pine oil saving method, a tert-butyl alcohol gum method, a dioxane gum method and the like. The method basically comprises the following steps: decolorizing, dehydrating, transparentizing, sealing, and the like. However, practical results show that the existing permanent microscope slide manufacturing method needs to accurately measure the type and concentration of the solution, and the using temperature and time of the solution, and once the solution is not measured, the problem of low quality of a sample is easily caused. For example: the specimen is not completely dried, and bubbles are easy to appear in the sealing piece, so that the surface of the worm is fuzzy and is not beneficial to observation; the problems of shrinkage and deformation of the body structure of the insect body due to improper control of the alcohol concentration in the dehydration process are easily caused by improper use of the alcohol concentration.

Disclosure of Invention

1. Technical problem to be solved

The technical problem to be solved by the invention is to provide a method for manufacturing permanent microscopic slide specimens of tiny insects, which simplifies the manufacturing process of microscopic slide specimens, achieves the control of specimen manufacturing quality by accurately measuring the type and concentration of solution, and the use temperature and time thereof, and avoids the problems of air bubbles in a sealing surface, shrinkage and deformation of an insect body structure in the dehydration process and the like.

2. Technical scheme

In order to achieve the purpose, the invention provides the following technical scheme: a method for manufacturing a permanent microscopic slide specimen of a tiny insect comprises the following steps:

step 1, soaking living insects in 65-76% alcohol solution;

step 2, taking out the insects from the alcohol solution, cutting the insects into bodies, putting the insects into a beaker added with a sodium hydroxide solution, putting the beaker on a heating plate, heating and soaking the insects, and cleaning the insects and the internal organs until the insects become transparent;

step 3, taking the polypide treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the polypide into constant-temperature cold bath acid fuchsin alcohol solution with gradually increased concentration for color fixation;

step 4, moving the polypide processed in the step 3 out, placing the polypide on filter paper, and placing the polypide in a constant-temperature vacuum chamber for drying and dehydration;

step 5, spraying xylene vapor to the inner and outer surfaces of the insects in the vacuum chamber in the step 4 to form a coating of 5-10 microns;

and 6, dripping Canadian gum on the glass slide, placing the insects treated in the step 5 on the Canadian gum, laying all parts of the insect body, covering a cover glass, and airing to obtain a permanent glass slide specimen.

In the method for preparing a microscope slide specimen, the concentration of the sodium hydroxide solution is 5%.

In the method for manufacturing the microscope slide specimen, in the step 2, the heating temperature of the heating plate is 100-120 ℃, and the heating time is 20 min.

In the method for preparing a microscope slide specimen, in step 3, the acid fuchsin alcohol solution uses alcohol having a concentration of 30%, 55%, 65%, 90%, 100% as a solvent.

In the method for preparing the microscope slide specimen, in step 3, the worm body is respectively soaked in the acid fuchsin alcohol solution with the alcohol solvent concentration of 30%, 55%, 65% and 90% for 40-80 min, and then soaked in the acid fuchsin alcohol solution with the alcohol solvent concentration of 100% for 80-160 min.

The preparation method of the microscope slide specimen is characterized in that the color fixing time in the step 3 is 4-8 h.

In the preparation method of the microscope slide specimen, the temperature of the constant-temperature vacuum chamber in the step 4 is 25 ℃, and the worm body is dried and dehydrated on the filter paper until no solvent exists on the filter paper.

3. Advantageous effects

In conclusion, the beneficial effects of the invention are as follows:

(1) compared with the existing specimen preparation method, the method for preparing the slide specimen has the advantages that the steps are simple, the used materials cannot cause environmental pollution, the dyeing effect is good, and the complexity of the specimen preparation process and the harmfulness to a human body can be reduced;

(2) according to the method for manufacturing the slide specimen, the specimen is dried through the constant-temperature vacuum chamber, the solution in the washing body is easy to fully evaporate, and the problems of internal bubbles caused by the existing natural air drying and further turbidity on the surface of the specimen are solved;

(3) according to the method for manufacturing the glass slide specimen, the polypide is dyed by the acid fuchsin alcohol solution, the body structure of the polypide can be more obviously displayed, and the shrinkage and deformation of the polypide structure can not be caused by the layer-by-layer dehydration of the alcohol with different concentrations.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

Example 1:

step 1, soaking living brown planthoppers in 65% alcohol solution;

step 2, taking the brown planthopper out of the alcohol solution, cutting the body, putting the body into a beaker added with 5% sodium hydroxide solution, placing the beaker on a heating plate, heating the beaker at 120 ℃, soaking the beaker for 20min, and cleaning the content and the viscera until the brown planthopper bodies become transparent;

step 3, taking the brown planthopper polypide treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the brown planthopper polypide into a constant-temperature cold-bath acid fuchsin alcoholic solution with alcoholic solvent concentration of 30%, 55%, 65% and 90% for 40min and a constant-temperature cold-bath acid fuchsin alcoholic solution with alcoholic solvent concentration of 100% for 100min to finish the color fixation of the brown planthopper polypide;

step 4, moving the brown planthopper polypide treated in the step 3 out, placing the brown planthopper polypide on filter paper, and placing the brown planthopper polypide in a vacuum chamber with constant temperature of 25 ℃ for drying and dehydrating until no solvent exists on the filter paper;

step 5, spraying xylene vapor to the inner and outer surfaces of the brown planthopper insects in the vacuum chamber in the step 4 to form a 6-micron coating;

and 6, dripping Canadian gum on the glass slide, placing the brown planthopper polypide treated in the step 5 on the Canadian gum, putting all parts of the brown planthopper polypide, covering a cover glass, and airing to obtain a permanent brown planthopper glass slide specimen I.

Example 2:

step 1, soaking a wheat aphid living body in 70% alcohol solution;

step 2, taking the wheat aphids out of the alcohol solution, cutting the bodies, putting the bodies into a beaker added with 5% sodium hydroxide solution, placing the beaker on a heating plate, heating the beaker at 100 ℃, soaking the beaker for 20min, and cleaning the inclusions and internal organs until the bodies of the insects become transparent;

step 3, taking the wheat aphid bodies treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the wheat aphid bodies into constant-temperature cold-bath acid fuchsin alcohol solutions with alcohol solvent concentrations of 30%, 55%, 65% and 90% for 50min and constant-temperature cold-bath acid fuchsin alcohol solutions with alcohol solvent concentrations of 100% for 120min to finish color fixation of the wheat aphid bodies;

step 4, moving the wheat aphid bodies treated in the step 3 out, placing the wheat aphid bodies on filter paper, and placing the filter paper in a vacuum chamber with constant temperature of 25 ℃ for drying and dehydrating until no solvent exists on the filter paper;

step 5, spraying xylene vapor to the inner and outer surfaces of the wheat aphids in the vacuum chamber in the step 4 to form a 6-micron coating;

and 6, dripping Canadian gum on the glass slide, placing the wheat aphid treated in the step 5 on the Canadian gum, laying each part of the wheat aphid, covering a cover glass, and drying to obtain a permanent glass slide specimen II.

Example 3:

step 1, soaking a living coccid in 76% alcohol solution;

step 2, taking out the scale insects from the alcohol solution, cutting the scale insects into bodies, putting the bodies into a beaker added with 5% sodium hydroxide solution, placing the beaker on a heating plate, heating the beaker at 100 ℃, soaking the beaker for 20min, and cleaning the content and the viscera until the scale insects become transparent;

step 3, taking the coccid bodies treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the coccid bodies into constant-temperature cold-bath acid fuchsin alcohol solutions with alcohol solvent concentrations of 30%, 55%, 65% and 90% for 45min and constant-temperature cold-bath acid fuchsin alcohol solutions with alcohol solvent concentrations of 100% for 100min to finish the color fixation of the coccid bodies;

step 4, the coccid bodies treated in the step 3 are moved out and placed on filter paper, and are placed in a vacuum chamber with constant temperature of 25 ℃ for drying and dehydration until no solvent exists on the filter paper;

step 5, spraying xylene vapor to the inner and outer surfaces of the coccid in the vacuum chamber in the step 4 to form a coating with the thickness of 5 microns;

and 6, dripping Canadian gum on the glass slide, placing the scale insects processed in the step 5 on the Canadian gum, laying all parts of the scale insects, covering with a cover glass, and drying to obtain a permanent glass slide specimen III.

Example 4:

step 1, placing a head louse living body in 65% alcohol solution for soaking;

step 2, taking out the head lice from the alcohol solution, cutting the head lice, putting the head lice into a beaker added with 5% of sodium hydroxide solution, placing the beaker on a heating plate, heating the beaker at 110 ℃, soaking the beaker for 20min, and cleaning the content and the viscera until the head lice bodies become transparent;

step 3, taking the head louse polypide treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the head louse polypide into constant-temperature cold-bath acid fuchsin alcohol solutions with alcohol solvent concentrations of 30%, 55%, 65% and 90% for 50min and constant-temperature cold-bath acid fuchsin alcohol solutions with alcohol solvent concentrations of 100% for 120min to finish color fixation of the head louse polypide;

step 4, moving the head lice bodies treated in the step 3 out, placing the head lice bodies on filter paper, and placing the head lice bodies in a vacuum chamber with constant temperature of 25 ℃ for drying and dehydrating until no solvent exists on the filter paper;

step 5, spraying xylene vapor to the inner and outer surfaces of the head lice in the vacuum chamber in the step 4 to form a coating with the thickness of 5 microns;

and 6, dripping Canadian gum on the glass slide, placing the head louse treated in the step 5 on the Canadian gum, laying all parts of the body of the head louse, covering a cover glass, and airing to obtain a permanent glass slide specimen IV.

Example 5:

step 1, soaking a pelteobagrus living body in 72% alcohol solution;

step 2, taking the pelagia pelteobagrus out of the alcohol solution, cutting the bodies, putting the pelagia pelteobagrus into a beaker with 5% sodium hydroxide solution, placing the beaker on a heating plate, heating the beaker at 100 ℃, soaking the beaker for 20min, and cleaning the content and the viscera until the pelagia pelteobagrus bodies become transparent;

step 3, taking the pelagia pelaginosum bodies treated in the step 2 out of the sodium hydroxide solution, and sequentially putting the pelagia pelaginosum bodies into a constant-temperature cold-bath acid fuchsin alcohol solution with alcohol solvent concentration of 30%, 55%, 65% and 90% for 45min and a constant-temperature cold-bath acid fuchsin alcohol solution with alcohol solvent concentration of 100% for 130min to finish color fixation of the pelagia pelaginosum bodies;

step 4, moving the pelagia round bodies processed in the step 3 out, placing the bodies on filter paper, and placing the bodies in a vacuum chamber with constant temperature of 25 ℃ for drying and dehydrating until no solvent exists on the filter paper;

step 5, spraying xylene vapor to the inner and outer surfaces of the pelecypodium pelecypomum in the vacuum chamber in the step 4 to form a coating with the thickness of 8 microns;

and 6, dripping Canadian gum on the glass slide, placing the pelecypris pelagi treated in the step 5 on the Canadian gum, laying all parts of the body of the pelecypris pelagi, covering a cover glass, and drying in the air to obtain a permanent glass slide specimen V.

Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.