CN102519774B - Dyeing method for peripheral nerves of fish larvae and juveniles - Google Patents
Dyeing method for peripheral nerves of fish larvae and juveniles Download PDFInfo
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- CN102519774B CN102519774B CN201110362535.XA CN201110362535A CN102519774B CN 102519774 B CN102519774 B CN 102519774B CN 201110362535 A CN201110362535 A CN 201110362535A CN 102519774 B CN102519774 B CN 102519774B
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- fish
- fish body
- alcohol
- body sample
- dyeing
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Abstract
The invention relates to a dyeing method for peripheral nerves of fish larvae and juveniles. The method comprises the following steps of: immersing a fish sample in 0.5 percent by weight of KOH solution and 3-4 percent by volume of H2O2 solution; performing alcohol dehydration processing and then immersing in 30-50 percent by weight of Sudan black solution to perform nerve dyeing; and performing alcohol hydration processing and then placing in 0.5 percent by weight of KOH. The method is easy to operate; the required reagent is less; the dyeing time is greatly shortened; expensive reagents and especial instruments are not required; and the method is easy to operate and stable in result, can be used for researching peripheral nerve distribution of normal fish larvae and juveniles and has a wider application range.
Description
Technical field
The invention belongs to the peripheroneural dyeing of fish field, particularly the peripheroneural colouring method of a kind of fish larvae juvenile fish.
Background technology
Nervous system is very important to the vital movement of fish, and it consists of brain, cranial nerve, spinal cord and spinal nerve, and brain and spinal cord are nervous centralis, and cranial nerve and spinal nerve are peripheral nerve; The research that fish peripheral neverous system distributes can provide important evidence for the research of fish origin and evolution and physiological function.Domestic there is not yet of research method report that relevant fish peripheral nerve distributes, external relevant report mainly concentrates on the following aspects: the neural decoration method of haematoxylin, the method Color is not good; Cartilage, os osseum, neural triple staining, main process first Yong Ali Xinlan dyes cartilage for blueness, secondly by sodium alizarinsulfonate, os osseum is dyed for redness, last peripheral nerve is after sudan black dyeing, aobvious black, Color is good, but this dyeing course is comparatively loaded down with trivial details.
Summary of the invention
Technical matters to be solved by this invention is to provide the peripheroneural colouring method of a kind of fish larvae juvenile fish, and the method reagent simple to operate, required is few, has greatly shortened dyeing time, reagent that also need not be expensive and special instrument.
The peripheroneural colouring method of a kind of fish larvae juvenile fish of the present invention, comprising:
(1) use fixedly fish body sample 48～54h of 10vol% formalin, the fish body sample fixing is rinsed to 12-24h;
(2) above-mentioned fish body is put into container, add 0.5wt%KOH solution until fish body is flooded, place 1-2h;
(3) to dripping concentration in above-mentioned solution, be 3～4vol%H
2o
2solution, is placed in container under incandescent lamp and spends the night, to accelerate to take off fish epidermis pigment;
(4) fish body is carried out to dehydration of alcohol processing, put into successively 30vol%, 50vol%, 70vol% alcohol, each 30～40min;
(5) fish body is immersed in 30wt%-50wt% sudan black solution and carry out nerve dyeing, dyeing time is 8-12h;
(6) fish body is carried out to alcohol hydration process, put into successively 70vol%, 60vol%, 50vol% alcohol, each 1-2min;
(7) fish body is placed in to 0.5wt%KOH solution and spends the night, complete peripheroneural dyeing.
10vol% formalin in described step (1) is diluted and is obtained by 37vol%-40vol% formalin.
Fish body sample in described step (1) is fish larvae juvenile fish sample.
Container in described step (2) is plastics or glass rectangular box.
In described step (3), every 100ml KOH dropping 4-6 drips H
2o
2.
Nerve dyeing in described step (5) is carried out in shaking table, and Color is better.
30wt%-50wt% sudan black solution preparation technique in described step (5) is: sudan black powder dissolution is in 70vol% alcohol, to state of saturation; With 70vol% alcohol, be diluted to 30wt%-50wt%.
In 0.5wt%KOH solution, slowly add glycerine, until fish body specimen samples sinks to bottom, place after 12h, remove gently head epidermis, under stereoscope, high-visible peripheral nerve distributes, and peripheral nerve dyes after treatment for black.
beneficial effect
(1) the present invention's reagent simple to operate, required is few, has greatly shortened dyeing time, reagent that also need not be expensive and special instrument;
(2) in the operation of the present invention, the soluble in water and glycerine of KOH, and the similar NaOH of its chemical property, have strong basicity, can destroy cell tissue; H
2o
2its aqueous solution is commonly called as hydrogen peroxide, and outward appearance is colourless transparent liquid, is a kind of strong oxidizer; KOH and H
2o
2be combined with and can make fish body skeleton softening, eliminate musculature, take off epidermis pigment, make fish body transparent, be convenient to observe fish body peripheral nerve dyeing situation; According to KOH, be soluble in the feature of glycerine, after dyeing, fish body is put into the mixed liquor of KOH and glycerine, not only makes that fish body is transparent to be convenient to neural dyeing and to observe, and can keep fish body moisture, and isolated air, is convenient to sample long preservation;
(3) the present invention is easy and simple to handle, and result is stable, can be used for the research that general fish larvae juvenile fish peripheral nerve distributes, and its scope of application is wider.
Embodiment
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment are only not used in and limit the scope of the invention for the present invention is described.In addition should be understood that those skilled in the art can make various changes or modifications the present invention after having read the content of the present invention's instruction, these equivalent form of values fall within the application's appended claims limited range equally.
Embodiment 1
(1) with fixing mandarin sturgeon prelarva sample (total length 6.2cm) 48h of 10vol% formalin (dilute and obtain by 37vol% formalin), the fish body sample fixing is rinsed to 12h;
(2) above-mentioned fish body is put into glass rectangular box, add 0.5wt%KOH solution until fish body is flooded, place 1h;
(3) to dripping concentration in above-mentioned solution, be 3vol%H
2o
2(100ml KOH drips 4～6 H to solution
2o
2), glass rectangular box is placed under incandescent lamp and is spent the night, to accelerate to take off fish epidermis pigment;
(4) fish body is carried out to dehydration of alcohol processing, put into successively 30vol%, 50vol%, 70vol% alcohol, each 30min;
(5) (sudan black powder dissolution is in 70vol% alcohol, to state of saturation fish body to be immersed to 30wt% sudan black solution; With 70vol% alcohol dilution) in carry out nerve and dye, dyeing time is 8h;
(6) fish body is carried out to alcohol hydration process, put into successively 70vol%, 60vol%, 50vol% alcohol, each 1min;
(7) fish body is placed in to 0.5wt%KOH solution and spends the night, complete peripheroneural dyeing.
Embodiment 2
(1) with the fixing fish body sample 54h of 10vol% formalin (dilute and obtain by 40vol% formalin), the fish body sample fixing is rinsed to 24h;
(2) above-mentioned fish body is put into plastics, add 0.5wt%KOH solution until fish body is flooded, place 2h;
(3) to dripping concentration in above-mentioned solution, be 4vol%H
2o
2(100ml KOH drips 4～6 H to solution
2o
2), plastics are placed under incandescent lamp and are spent the night, to accelerate to take off fish epidermis pigment;
(4) fish body is carried out to dehydration of alcohol processing, put into successively 30vol%, 50vol%, 70vol% alcohol, each 40min;
(5) (sudan black powder dissolution is in 70vol% alcohol, to state of saturation fish body to be immersed to 50wt% sudan black solution; With 70vol% alcohol dilution) in shaking table, carry out nerve and dye, dyeing time is 12h;
(6) fish body is carried out to alcohol hydration process, put into successively 70vol%, 60vol%, 50vol% alcohol, each 2min;
(7) fish body is placed in to 0.5wt%KOH solution and spends the night, complete peripheroneural dyeing.
Claims (5)
1. the peripheroneural colouring method of fish larvae juvenile fish, comprising:
(1) use fixedly fish body sample 48～54h of 10vol% formalin, the fish body sample fixing is rinsed to 12-24h; Wherein, 10vol% formalin is diluted and is obtained by 37vol%-40vol% formalin; Wherein, fish body sample is fish larvae juvenile fish sample;
(2) above-mentioned fish body sample is put into container, add 0.5wt%KOH solution until fish body sample is flooded, place 1-2h;
(3) to dripping concentration in above-mentioned solution, be 3～4vol%H
2o
2solution, is placed in container under incandescent lamp and spends the night;
(4) fish body sample is carried out to dehydration of alcohol processing, put into successively 30vol%, 50vol%, 70vol% alcohol, each 30～40min;
(5) fish body sample is immersed in 30wt%-50wt% sudan black solution and carry out nerve dyeing, dyeing time is 8-12h;
(6) fish body sample is carried out to alcohol hydration process, put into successively 70vol%, 60vol%, 50vol% alcohol, each 1-2min;
(7) fish body sample is placed in to 0.5wt%KOH solution and spends the night, complete peripheroneural dyeing.
2. the peripheroneural colouring method of a kind of fish larvae juvenile fish according to claim 1, is characterized in that: the container in described step (2) is plastics or glass rectangular box.
3. the peripheroneural colouring method of a kind of fish larvae juvenile fish according to claim 1, is characterized in that: in described step (3), every 100ml KOH dropping 4-6 drips H
2o
2.
4. the peripheroneural colouring method of a kind of fish larvae juvenile fish according to claim 1, is characterized in that: the nerve dyeing in described step (5) is carried out in shaking table.
5. the peripheroneural colouring method of a kind of fish larvae juvenile fish according to claim 1, is characterized in that: the 30wt%-50wt% sudan black solution preparation technique in described step (5) is: sudan black powder dissolution is in 70vol% alcohol, to state of saturation; With 70vol% alcohol, be diluted to 30wt%-50wt%.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110362535.XA CN102519774B (en) | 2011-11-16 | 2011-11-16 | Dyeing method for peripheral nerves of fish larvae and juveniles |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110362535.XA CN102519774B (en) | 2011-11-16 | 2011-11-16 | Dyeing method for peripheral nerves of fish larvae and juveniles |
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| Publication Number | Publication Date |
|---|---|
| CN102519774A CN102519774A (en) | 2012-06-27 |
| CN102519774B true CN102519774B (en) | 2014-02-12 |
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| CN201110362535.XA Expired - Fee Related CN102519774B (en) | 2011-11-16 | 2011-11-16 | Dyeing method for peripheral nerves of fish larvae and juveniles |
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Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104596825A (en) * | 2014-12-26 | 2015-05-06 | 中国水产科学研究院南海水产研究所 | Dyeing method of fish bones |
| CN105987838A (en) * | 2015-02-06 | 2016-10-05 | 中国科学院上海生命科学研究院 | Tissue section visualized processing method |
| CN110495445A (en) * | 2019-09-20 | 2019-11-26 | 河南师范大学 | A kind of method of the Yellow River carp postlarva intermuscular bone dyeing |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1961816A (en) * | 2006-11-24 | 2007-05-16 | 山东建筑大学 | Method for making fetus transparent specimen |
| CN101915693A (en) * | 2010-07-07 | 2010-12-15 | 新疆医科大学 | Human embryonic trigeminus based three-dimensional reconstruction method by using histotomy staining |
-
2011
- 2011-11-16 CN CN201110362535.XA patent/CN102519774B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1961816A (en) * | 2006-11-24 | 2007-05-16 | 山东建筑大学 | Method for making fetus transparent specimen |
| CN101915693A (en) * | 2010-07-07 | 2010-12-15 | 新疆医科大学 | Human embryonic trigeminus based three-dimensional reconstruction method by using histotomy staining |
Non-Patent Citations (6)
| Title |
|---|
| Clearing and Staining Whole Fish Specimens for Simultaneous Demonstration of Bone, Cartilage, and Nerves;Jiakun Song等;《Copeia》;19950215;第1995卷(第1期);第115-116页 * |
| Jiakun Song等.Clearing and Staining Whole Fish Specimens for Simultaneous Demonstration of Bone, Cartilage, and Nerves.《Copeia》.1995,第1995卷(第1期),115-116. |
| Sihler肌内神经染色方法的探讨;李平等;《遵义医学院学报》;20010831;第24卷(第4期);第317-319页 * |
| 张倩等.点纹斑竹鲨晚期胚胎侧线系统的研究.《上海海洋大学学报》.2011,第20卷(第1期),第76-79页. |
| 李平等.Sihler肌内神经染色方法的探讨.《遵义医学院学报》.2001,第24卷(第4期),第317-319页. |
| 点纹斑竹鲨晚期胚胎侧线系统的研究;张倩等;《上海海洋大学学报》;20110131;第20卷(第1期);第76-79页 * |
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Address after: 200090 Shanghai military road, Yangpu District, No. 300 Applicant after: East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences Address before: 200090 No. 100, military road, Shanghai, Yangpu District Applicant before: East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences |
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