CN104764645A - Amphibian phalanx paraffin section manufacturing method - Google Patents
Amphibian phalanx paraffin section manufacturing method Download PDFInfo
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- CN104764645A CN104764645A CN201510208343.1A CN201510208343A CN104764645A CN 104764645 A CN104764645 A CN 104764645A CN 201510208343 A CN201510208343 A CN 201510208343A CN 104764645 A CN104764645 A CN 104764645A
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Abstract
The invention relates to an animal bone processing method, in particular to an amphibian phalanx paraffin section manufacturing method. Amphibian phalanxes are fixed and stored through a 10% physiological salt formaldehyde solution, decalcification is carried out, and then the decalcified phalanxes are dyed, dehydrated, subjected to hyalinizing, embedded, sliced, attached and packaged by means of Harris hematoxylin staining liquid through a fading dyeing method. The method is easy to operate, short in consumed time, good in dyeing effect, suitable for meeting the requirement for amphibian phalanx paraffin section manufacturing on a large scale in scientific research and wide animal protection work, and capable of improving accuracy of population age assessment.
Description
Technical field
The present invention relates to a kind of method to animal skeleton process, be specially amphibian animal phalanx paraffin section method for making.
Background technology
Age composition is the important foundation of postgraduate's species group number change, has great importance for the rescue of the control of harmful animal, the conservation and utilization of wildlife resources and animals on the brink of extinction population and recovery.The growth of bone is relevant to nutrition condition, metaboilic level, upgrowth situation, envirment factor etc., thus bone tissue density degree is differed, and presents torus feature.Therefore, the torus of bone is important materials and the foundation of animal age qualification, as the torus of vertebrae joint concave surface.
Research finds that amphibian animal phalanx also presents torus feature, and it is consistent that its age and the ring on phalanx take turns number.Utilize the ring on phalanx to take turns number and farthest can reduce injury to animal to carry out amphibian Age Determination, realize the sustainable use of Animal resources.Therefore, utilize toe bone tissue paraffin section to differentiate that the method at amphibian animal age is generally used.Usual toe bone tissue paraffin section method complex steps, length consuming time, complicated operation, and Color is difficult to be guaranteed, and directly has influence on the judgement of amphibian animal population age.
Summary of the invention
The invention provides one for above-mentioned technical matters, operation steps simple amphibian animal phalanx paraffin section method for making, concrete technical scheme is:
Amphibian animal phalanx paraffin section method for making, comprises the following steps:
(1) preserve fixing for the amphibian animal phalanx removing appearance skin and musculature by the physiology salt formalin of 10%, the time is 2-4 hour;
(2) by the physiology salt salpeter solution of 5-10% or mass concentration be the aqueous solution of nitric acid of 5-10% by the amphibian animal phalanx decalcification after fixing, the time is 24-48 hour;
(3) with Harris brazilwood extract dyeing liquid by the phalanx bulk dyeing after decalcification 140 minutes, colouring method is for taking off back decoration method;
(4) then routinely means phalanx is dewatered, transparent, embedding, section, paster, mounting process.
Described 10% physiology salt formalin, the formalin for every 10ml adds 90ml physiological saline, and formalin concentration is 37-40%, i.e. formalin solution, and physiological saline is mass concentration 0.65% sodium chloride solution.
The mass concentration of described Harris brazilwood extract dyeing liquid is 0.5%.
Described amphibian animal phalanx is hind leg the 4th toe, i.e. the longest toe.
Amphibian animal phalanx paraffin section method for making provided by the present invention, by the step that simplifies the operation, optimize each step Production Time, adjustment staining procedure, adopt take off back colouring method, raising Color.This method overcomes the complex steps of classic method, length consuming time, complicated operation, and Color is difficult to be guaranteed, and directly has influence on the problems such as the accuracy of amphibian animal population age judgement.This method is simple to operate, consuming time less, Color is good, to be adapted in scientific research and conservation of wildlife work, to the requirement that the amphibian animal phalanx paraffin section of extensive, mass makes, improve the accuracy rate of population age qualification.
Accompanying drawing explanation
Fig. 1 is the high eyebrow tree frog amphibian animal phalanx paraffin section observation figure that classic method obtains;
Fig. 2 is the present embodiment high eyebrow tree frog amphibian animal phalanx paraffin section observation figure.
Embodiment
The invention will be further described with case study on implementation by reference to the accompanying drawings.
(1) draw materials, fix:
Get the 4th toe of high eyebrow tree frog, the skin of removing outside and musculature, fix 2-4 hour by 10% physiology salt formalin;
(2) decalcification, namely soften:
With the physiology salt salpeter solution of 5-10% or the aqueous solution of nitric acid of 5-10% by the sample decalcification after fixing, the time is about 24-48 hour, till bone tissue is softening, can thrust bone tissue gently with pin;
(3) dye, whole dye method:
With Harris brazilwood extract dyeing, the sample after decalcification is carried out bulk dyeing 140 minutes, colouring method is for taking off back decoration method, feature immerses in the higher dye liquor of concentration by by dye material, make its overstain, and then carry out fading or color separation with chemical agent, dyestuff unnecessary in removing tissue, reaches required colourity.Taking off back in decoration method, in order to color of cutting into slices afterwards in decolouring obtains distincter differentiation, the overstain time generally extends.
(4) flowing water oil blackeite:
By the sample flowing water oil blackeite that dyeed 30 minutes.
(5) dewater:
Ethanolic solution sample good for oil blackeite first being put into respectively 70% and 80% respectively dehydration one hour, then put into 1% eosin alcoholic solution and contaminate 5-10 minute, 1% eosin alcoholic solution is that every 1 gram of eosin adds in the ethanolic solution of 100 milliliter 95% formulated; And then dewater one hour with the ethanolic solution of 95%, unnecessary redness can be washed away simultaneously, dewater one hour finally by anhydrous alcohol.
(6) transparent:
With transparent 15 minutes of the mixed liquor of dimethylbenzene and alcohol 1: 1, put into pure dimethylbenzene transparent 30 minutes, change a solution therebetween.
(7) waxdip:
This step is carried out in constant temperature oven, transparent good sample is immersed in the mixed liquor of dimethylbenzene and paraffin 1: 1, the constant temperature oven be placed in higher than paraffin melting point 2 degree keeps half an hour, is then moved into by frog phalanx and waxdip is housed in the wax cup of paraffin refined wax one hour, change a wax cup therebetween.
(8) embed:
When embedding the paraffin in carton and being in half curdled appearance, immediately the phalanx having soaked wax is placed vertically in a certain order in carton, the also submergence phalanx completely of the wax now in carton, should continue to add the wax that dissolves immediately until submergence phalanx.
(9) cut into slices
Embedded sample is cut into the thin slice that thickness is 12-13 μm, and slice thickness becomes positive correlation with dyeing time with the color separation time, and thin section is more of light color relative to slab.
(10) paster and exhibition sheet
A. during paster, microslide is coated a small amount of alite paste, adding a little distilled water in being coated with flat alite paste surface, making it evenly be shattered into skim;
B. wax band is divided into suitable segment or little wax disk(-sc), shiny surface is downward, is arranged on the distillation water layer on sticky posts agent surface;
C. steadily picked up by microslide, slightly heat below with spirit lamp, wax band segment flattens because being heated;
D. open up after sheet, suitably tilt, allow unnecessary distilled water diffluence, or sop up with thieving paper.
(11) roasting sheet:
Be placed in the constant temperature oven of 37 to 40 DEG C by the microslide posting wax band, roasting sheet 5-10 hour, wax disk(-sc) is attached on microslide securely.
(12) dewax
The paraffin section dried is placed in the container filling dimethylbenzene, dissolves away the paraffin on it, need 30-40 minute, time length and temperature correlation.
Fig. 1 is the high eyebrow tree frog amphibian animal phalanx paraffin section observation figure that classic method obtains, Fig. 2 is the present embodiment high eyebrow tree frog amphibian animal phalanx paraffin section observation figure, contrast as can be seen from two width figure, wheel loop wire in the section bone that the present embodiment obtains is high-visible, improves the accuracy rate of Age Determination; And film-making time shorten, efficiency improves.
Claims (3)
1. amphibian animal phalanx paraffin section method for making, is characterized in that, comprise the following steps:
(1) preserve fixing for the amphibian animal phalanx removing appearance skin and musculature by the physiology salt formalin of 10%, the time is 2-4 hour;
(2) by the physiology salt salpeter solution of 5-10% or mass concentration be the aqueous solution of nitric acid of 5-10% by the amphibian animal phalanx decalcification after fixing, the time is 24-48 hour;
(3) dyeed 140 minutes by the phalanx after decalcification with Harris brazilwood extract dyeing liquid, colouring method is for taking off back decoration method;
(4) then routinely means phalanx is dewatered, transparent, embedding, section, paster, mounting process.
2. amphibian animal phalanx paraffin section method for making according to claim 1, it is characterized in that, 10% physiology salt formalin described in step (1), the formalin for every 10ml adds 90ml physiological saline, and physiological saline is mass concentration 0.65% sodium chloride solution.
3. amphibian animal phalanx paraffin section method for making according to claim 1 or 2, is characterized in that, the mass concentration of the Harris brazilwood extract dyeing liquid described in step (3) is 0.5%.
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| CN201510208343.1A CN104764645A (en) | 2015-04-27 | 2015-04-27 | Amphibian phalanx paraffin section manufacturing method |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105136548A (en) * | 2015-09-11 | 2015-12-09 | 安徽省农业科学院茶叶研究所 | Preparation method for paraffin section of ectropis obliqua male adult internal genitalia and tissue picking and moving tool for application of preparation method |
| CN108918215A (en) * | 2018-05-17 | 2018-11-30 | 苏州大学 | Quickly prepare the method and its application of hard tissue slicing |
| CN112396924A (en) * | 2020-12-04 | 2021-02-23 | 南方医科大学 | Method for embedding skull |
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| CN1083212A (en) * | 1993-06-08 | 1994-03-02 | 山西大学 | The easy quick paraffin sections method of human body and animal tissue |
| CN101042317A (en) * | 2007-04-27 | 2007-09-26 | 孝感学院 | Section staining method for striped skeletal muscles |
| CN103759994A (en) * | 2014-01-02 | 2014-04-30 | 河南科技大学 | Larva paraffin section forming method |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1083212A (en) * | 1993-06-08 | 1994-03-02 | 山西大学 | The easy quick paraffin sections method of human body and animal tissue |
| CN101042317A (en) * | 2007-04-27 | 2007-09-26 | 孝感学院 | Section staining method for striped skeletal muscles |
| CN103759994A (en) * | 2014-01-02 | 2014-04-30 | 河南科技大学 | Larva paraffin section forming method |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105136548A (en) * | 2015-09-11 | 2015-12-09 | 安徽省农业科学院茶叶研究所 | Preparation method for paraffin section of ectropis obliqua male adult internal genitalia and tissue picking and moving tool for application of preparation method |
| CN105136548B (en) * | 2015-09-11 | 2017-12-22 | 安徽省农业科学院茶叶研究所 | A kind of tissue of preparation method and application this method of tea geometrid male imago internal genitalia paraffin section chooses shifting instrument |
| CN108918215A (en) * | 2018-05-17 | 2018-11-30 | 苏州大学 | Quickly prepare the method and its application of hard tissue slicing |
| CN108918215B (en) * | 2018-05-17 | 2021-03-23 | 苏州大学 | Method for rapidly preparing hard tissue slices and application thereof |
| CN112396924A (en) * | 2020-12-04 | 2021-02-23 | 南方医科大学 | Method for embedding skull |
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