CN109060468A - A kind of preparation method of the paraffin section of pear flower tissue - Google Patents
A kind of preparation method of the paraffin section of pear flower tissue Download PDFInfo
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
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Abstract
The present invention relates to biological histology's technical fields, more particularly to a kind of preparation method of the paraffin section of pear flower tissue, include the following steps: to take out the pear flower saved in 4-5 degrees Celsius of refrigerator, it is clean with distilled water flushing, 3mm below pistil, stamen, calyx, petal and anthocaulus base portion is all removed, treated pear flower tissue is obtained；The pear flower tissue of acquisition is put into the FAA fixer of fixed container, fixed container is evacuated, until no longer there is bubble in FAA fixer, it is 1-2 days fixed after pumping；The pear flower tissue obtained after pumping is placed on gauze and blots FAA fixer, and is successively dehydrated, is transparent, the processing of waxdip, embedding, slice, bonding die, dewaxing, dyeing and removal floating color.Method provided by the present invention solves prior paraffin slice and is not strict in terms of material processing, and tissue is too big when the later period being caused to be sliced, and the defect of extreme influence is generated to experimental result, and obtained paraffin section quality is higher.
Description
Technical field
The present invention relates to biological histology's technical field more particularly to a kind of plant tissue paraffin section preparation method, tools
Body is related to a kind of preparation method of the paraffin section of pear flower tissue.
Background technique
Paraffin tissue sections technology is a kind of method most widely used in histology conven tional tabletting techniques.It is applied not only to
The morphosis and the subjects such as pathology and medical jurisprudence for observing normal cell tissue are to study, observe and judge groups of cells
The main method for the metamorphosis knitted is widely used in the research of numerous ambits.But due to the institutional framework of plant
The concrete operation method of specificity, different materials is also not quite similar.
Currently, the paraffin section technical research carried out on pear flower is less and the effect is unsatisfactory, it is complicated for operation.Therefore such as
The problem of what overcome the deficiencies in the prior art is current biological histology's technical field urgent need to resolve.
Summary of the invention
In order to solve the above technical problems, The present invention provides a kind of preparation methods of the paraffin section of pear flower tissue.
The present invention includes following technical scheme: a kind of preparation method of the paraffin section of pear flower tissue includes the following steps:
(1) pear flower saved in 4-5 degrees Celsius of refrigerator is taken out, it is clean with distilled water flushing, by pistil, stamen, flower
3mm below calyx, petal and anthocaulus base portion is all removed, and obtains treated pear flower tissue；
Material processing is the first step of paraffin section, how to handle material and directly influences subsequent experiment and final paraffin
The quality of piece.Traditional paraffin section does not clearly require in terms of material processing, it is only necessary to which it is solid that size can be immersed in FAA
Determine in liquid.Since prior paraffin slice is not strict in terms of material processing, tissue is too big when the later period being caused to be sliced,
It is extremely inconvenient in finishing operations, extreme influence is generated to paraffin section quality.Therefore in the present invention in pear flower tissue treatment,
Extra tissue is removed.
(2) the pear flower tissue obtained in step (1) is put into the FAA fixer of fixed container, fixed container is carried out
Pumping, until no longer there is bubble in FAA fixer, it is 1-2 days fixed after pumping；
Material is only placed in fixer and is fixed by traditional fixation procedure, but has ignored space between cells in material
Air in later period embedding process, can be discharged due to heated if these air are not excluded for, lead to material table in embedding wax block
There is bubble in face, influences later period slice.The present invention is extracted out air using vacuum pump, can dramatically remove space between cells
Air, improve the later period embed quality.
(3) the pear flower tissue obtained after pumping is placed on gauze and blots FAA fixer, and be successively dehydrated, thoroughly
Bright, waxdip, embedding, slice, bonding die, dewaxing, dyeing and removal floating color processing.
Further, in the step (3) the step of dehydration are as follows: the pear flower group after FAA fixer will be blotted and be woven in
According to 70%, 80%, 85%, 90%, 95% concentration of alcohol, respectively dehydration is primary step by step in cryopreservation tube, and each 2h is then used
95% dehydration of alcohol 1h, is repeated twice, and last dehydrated alcohol is dehydrated twice, each each 45min.
Further, step transparent in the step (3) are as follows: after the completion of dehydration, according to 1/2 countless+1/2 diformazans of ethyl alcohol
Benzene, pure dimethylbenzene sequence sample is reprocessed twice, each clearing time be 1.5h.
Further, the step of waxdip is handled in the step (3) are as follows:
Oven temperature is adjusted to 80 degrees Celsius, is put into paraffin melting spare；
Prepare cryopreservation tube, after 1/2 dimethylbenzene is added, is put into baking oven, the paraffin of thawing is added in cryopreservation tube after 30 seconds,
Cryopreservation tube lid is sealed, is placed in 60 degrees Celsius of baking ovens；
From baking oven take out cryopreservation tube open, be put into complete it is transparent after material, sealing, continue to be put into 60 degrees Celsius of baking ovens
The middle time is 16h；
Material taking-up is put into pure wax liquor, at 60 c, every 2h is changed once, and replacement is twice.
Further, in the step (3) the step of embedding treatment are as follows: fold imbedded mold using brown paper；
Water-bath is set as 80 degrees Celsius, and beaker is placed in water-bath and is preheated, and it is pure that 80% is then added in beaker
Wax and 20% old wax；One block of cardboard is placed as station on water-bath top, and embedded box is placed on cardboard, half is poured into
Material is taken out from baking oven, is put into embedded box by paraffin；
Material angle is adjusted using tweezers, with outside at calyx, apart from paper wall 1cm, two materials are at a distance of 2cm；Adjustment terminates
After pour into other half paraffin, embedded box is placed on cooling, be cooled to upper layer occur a tunic when continuously add stone
Wax is to highest point.
The process of the step (3) is referred to as to mend wax, and embedding is one of the committed step of paraffin section production, if embedding matter
Measure it is poor, that it is subsequent operation can not almost carry out.Traditional embedding process, only material is packed in paraffin by simple introduce,
The case where unavoidable bubble, layering and the material for occurring generating after embedding can not be embedded all.Paraffin be it is noncrystal,
Its type of cooling and crystal be not identical, if only once falling wax, wax stone surface will appear a dell after cooling down, and can incite somebody to action when serious
Material exposes；If can not control temperature in embedding, in two times plus the process of wax will certainly generate lamination；It is fixed
When, if the emptying of sending down abnormally ascending bubble, later period can not generate a large amount of bubbles on the surface of the material, influence to be sliced.The present invention asks three above
Operating process is placed on water-bath by mending wax and vacuum pump processing is addressed by topic.Simultaneously we embedding when simultaneously
It is non-to use pure wax, but be formulated using pure wax and old wax, such wax has compared to traditional pure wax to extend well
Property, there is good connection function when being sliced to the later period.
Further, in the step (3) the step of slice are as follows:
Embedded box is opened, is trimmed to paraffin mass using pocket knife trapezoidal, is bottom at calyx, is upper bottom at anthocaulus, cuts
Firmly uniform when piece, notch is vertical, and small trapezoidal each face is smooth, and 1cm paraffin is reserved at anthocaulus, and paraffin is as few as possible at calyx,
It is remaining to reserve 2mm paraffin everywhere；
The paraffin reserved at anthocaulus is sticked on wood particle, guarantees that entire material is vertical with adhesive surface, is then fixed on hand
On the rotatable slicer of driving wheel, setting slice thickness is 10 microns, keeps vertical with blade at calyx；
In slicing processes, since pear flower tissue is larger, and each position size is uneven, and the dynamics for shaking slice handle is appropriate,
Turned down rapidly from top to when incision, from bottom to top when slow but speed it is uniform.
When being sliced using slicer, traditional dicing method requires revolving speed uniformly firmly consistent, but this
Slicing mode, just for smaller, the good situation of wax ductility of tissue.And pear flower tissue is after treatment, relative to other
Tissue is still bigger, if be at the uniform velocity sliced, can not obtain complete histotomy, and in cutter half, slice begins to roll up
Song can not go on.The method applied in the present invention is cut by up to ten thousand times shake, and is summarized, and effect is very good.
Further, bonding die step in the step (3) are as follows:
Use distilled water flushing glass slide, baking oven drying；Bonding die agent is obtained by filtration in egg white, is placed in 4 degrees Celsius of refrigerators
Middle preservation；
Bonding die agent is applied on glass slide, while adding 6-8 drop distilled water, it is applied using glass bar, guarantee on glass slide
There is bonding die agent, downward by the reverse side of paraffin section at this time, is placed in bonding die agent；
After placing, toasted using alcolhol burner in glass slide lower part, the paraffin around discovery slice disappears, and is sure not
Over-heating, so that tissue is destroyed；
Distilled water is sucked using filter paper after baking piece, group is woven in open and flat on glass slide at this time, and paraffin is covered on tissue surface, and
Tissue is tightly combined with glass slide.
The production of glass slide is the final step and one of the most key step of microsection manufacture, if effect is bad,
Later observations effect can be seriously affected.Traditional method for adhering piece is that egg white is applied on slide, after it has been done, is just added
Add distilled water, toasted on alcolhol burner, and the wax around tissue can not be melted.Although cell is cut in processing in this way
Piece has preferable effect, but for organizing greatly, can not make its slice flattening.The present invention can be by the complete exhibition of tissue
It is flat, even the slice of especially fold can also flatten, while making later period dewaxing effect more preferable compared to traditional approach.
Further, in the step (3) the step of dewaxing are as follows: glass slide is put into pure dimethylbenzene, dewaxing 4h, 3h,
2h；After guaranteeing that dewaxing effect, dimethylbenzene use 4-5 times, need replacing.
Further, in the step (3) the step of dyeing are as follows:
After the completion of dewaxing, slice is repeated twice according to+1/2 dehydrated alcohol of 1/2 paraffin, without watery wine, 95% alcohol repeats
Twice, the sequence progress rehydration processing of 90% alcohol, 85% alcohol, 80% alcohol, 70% alcohol, 50% alcohol, 3 points every time
Clock；
Glass slide after rehydration is placed on 1% sarranine solution dyeing 16h, glass slide is taken out and is placed in 85% alcohol and impregnate
It 5 seconds, is subsequently placed into 0.5% fast green solution and dyes 5 seconds.
Further, in the step (3) the step of removal floating color are as follows: after the completion of dyeing, glass slide is put in 90% wine
Essence, 95% alcohol is repeated twice, dehydrated alcohol is repeated twice middle removal floating color, and 15 seconds every time；Glass slide is put into pure dimethylbenzene
In it is transparent can observe pear flower tissue microstructure under an optical microscope after ten minutes, can clearly find, be dyed to red
The bast of xylem and green.
By adopting the above technical scheme, including it is following the utility model has the advantages that pear flower tissue provided by the present invention paraffin section
Preparation method solves prior paraffin slice and is not strict in terms of material processing, and tissue is too big when the later period being caused to be sliced,
It is extremely inconvenient in finishing operations, the defect of extreme influence is generated to experimental result, and processing is exhausted to material, it is made
The paraffin section quality obtained is higher.
Detailed description of the invention
Fig. 1 is vascular bundle distribution schematic diagram at anthocaulus；
Fig. 2 is single vascular bundle schematic diagram at anthocaulus；
Fig. 3 is vascular bundle distribution schematic diagram at calyx；
Fig. 4 is single vascular bundle schematic diagram at calyx.
Specific embodiment
The present invention is described in further detail below by specific embodiment.
Embodiment:
The preparation method for present embodiments providing a kind of paraffin section of pear flower tissue, includes the following steps:
S1, the pear flower saved in 4-5 degrees Celsius of refrigerator is taken out, it is clean with distilled water flushing, by pistil, stamen, flower
3mm below calyx, petal and anthocaulus base portion is all removed, and obtains treated pear flower tissue；
S2, the pear flower tissue obtained in step S1 is put into the FAA fixer of fixed container, fixed container is taken out
Gas, until no longer there is bubble in FAA fixer, it is 1-2 days fixed after pumping；
S3, the pear flower tissue obtained after pumping is placed on gauze and blots FAA fixer, and be successively dehydrated, thoroughly
Bright, waxdip, embedding, slice, bonding die, dewaxing, dyeing and removal floating color processing.
In the present embodiment, the step S3 is specially；
S301 dehydration: will blot the pear flower group after FAA fixer be woven in cryopreservation tube according to 70%, 80%, 85%,
90%, respectively dehydration is primary step by step for 95% concentration of alcohol, and each 2h then uses 95% dehydration of alcohol 1h, is repeated twice, last nothing
Water-ethanol is dehydrated twice, each each 45min.
Further, step transparent in the step (3) are as follows: after the completion of dehydration, according to 1/2 countless+1/2 diformazans of ethyl alcohol
Benzene, pure dimethylbenzene sequence sample is reprocessed twice, each clearing time be 1.5h.
The processing of S302 waxdip: oven temperature is adjusted to 80 degrees Celsius, is put into paraffin melting spare；
Prepare cryopreservation tube, after 1/2 dimethylbenzene is added, is put into baking oven, the paraffin of thawing is added in cryopreservation tube after 30 seconds,
Cryopreservation tube lid is sealed, is placed in 60 degrees Celsius of baking ovens；
From baking oven take out cryopreservation tube open, be put into complete it is transparent after material, sealing, continue to be put into 60 degrees Celsius of baking ovens
The middle time is 16h；
Material taking-up is put into pure wax liquor, at 60 c, every 2h is changed once, and replacement is twice.
S303 embedding treatment: imbedded mold is folded using brown paper；Mold size foundation holds obtained paraffin section
Number determine, if two materials, long 5cm, wide 1cm, high 1.5cm mold can be folded.
Water-bath is set as 80 degrees Celsius, and beaker is placed in water-bath and is preheated, and it is pure that 80% is then added in beaker
Wax and 20% old wax；Old wax can be made with the following method, melt again after the cooling of 3 melted paraxylenes is added in the pure wax of 100ml,
It can be prepared by repeatedly for three times, be placed in beaker and melt, paraffin solidifies very much fastly when embedding in order to prevent.
One block of cardboard is placed as station on water-bath top, and embedded box is placed on cardboard, half paraffin is poured into, it will
Material takes out from baking oven, is put into embedded box；
Material angle is adjusted using tweezers, with outside at calyx, apart from paper wall 1cm, two materials are at a distance of 2cm；Adjustment terminates
After pour into other half paraffin, embedded box is placed on cooling, be cooled to upper layer occur a tunic when continuously add stone
Wax is to highest point.
It is sliced in S304: embedded box is opened, be trimmed to paraffin mass using pocket knife trapezoidal, be bottom, anthocaulus at calyx
Place is upper bottom, and when slice is firmly uniform, and notch is vertical, otherwise paraffin mass is easy to cause to shred；Small trapezoidal each face is smooth, at anthocaulus
1cm paraffin is reserved, paraffin is as few as possible at calyx, remaining reserves 2mm paraffin everywhere；
The paraffin reserved at anthocaulus is sticked on wood particle, guarantees that entire material is vertical with adhesive surface, is then fixed on hand
On the rotatable slicer of driving wheel, setting slice thickness is 10 microns, keeps vertical with blade at calyx；
In slicing processes, since pear flower tissue is larger, and each position size is uneven, and the dynamics for shaking slice handle is appropriate,
Turned down rapidly from top to when incision, from bottom to top when slow but speed it is uniform, can just cut out suitable slice, this process test cut
Piece person is patient and paraffin component, otherwise will lead to that paraffin does not form and gauffer even paraffin is broken；Just starting slice is paraffin,
It will appear histotomy below, since angle adjustment is improper, the biopsy tissues for just starting to occur are not completed, after organizing completely
(can visually observe), gently being pricked using dissecting needle in wax disk(-sc) blank space can remove, and make sure to keep in mind to distinguish slice front and back sides；
S305 bonding die: distilled water flushing glass slide, baking oven drying are used；Bonding die agent is obtained by filtration in egg white, is placed in 4 and takes the photograph
It is saved in family name's degree refrigerator；
Bonding die agent is applied on glass slide, while adding 6-8 drop distilled water, it is applied using glass bar, guarantee on glass slide
There is bonding die agent, downward by the reverse side of paraffin section at this time, be placed in bonding die agent, if counter put can find that slice can not be sticked to
On slide；
After placing, toasted using alcolhol burner in glass slide lower part, the paraffin around discovery slice disappears, and is sure not
Over-heating, so that tissue is destroyed；
Distilled water is sucked using filter paper after baking piece, group is woven in open and flat on glass slide at this time, and paraffin is covered on tissue surface, and
Tissue is tightly combined with glass slide.
S306 dewaxing: glass slide is put into pure dimethylbenzene, and dewax 4h, 3h, 2h；In order to guarantee dewaxing effect, dimethylbenzene
Using after 4-5 times, need replacing.
S307 dyeing: after the completion of dewaxing, will slice be repeated twice according to+1/2 dehydrated alcohol of 1/2 paraffin, without watery wine, 95%
Alcohol is repeated twice, the sequence of 90% alcohol, 85% alcohol, 80% alcohol, 70% alcohol, 50% alcohol carries out rehydration processing,
3 minutes every time；
Glass slide after rehydration is placed on 1% sarranine solution dyeing 16h, glass slide is taken out and is placed in 85% alcohol and impregnate
It 5 seconds, is subsequently placed into 0.5% fast green solution and dyes 5 seconds.
S308 removal floating color: after the completion of dyeing, glass slide is put in 90% alcohol, 95% alcohol is repeated twice, dehydrated alcohol
It is repeated twice middle removal floating color, 15 seconds every time；By glass slide be put into pure dimethylbenzene it is transparent after ten minutes, can be in optical microphotograph
Microscopic observation pear flower tissue microstructure, can clearly find, be dyed to the bast of red xylem and green.
Refering to fig. 1-4, the pear flower tissue vascular bundle paraffin section made with the present embodiment, can clearly observe pear flower group
Knit any portion of Vascular Bundle Characters.
The foregoing is only a preferred embodiment of the present invention, is not intended to restrict the invention, for the skill of this field
For art personnel, the invention may be variously modified and varied.All within the spirits and principles of the present invention, made any to repair
Change, equivalent replacement, improvement etc., should all be included in the protection scope of the present invention.
Claims (10)
1. a kind of preparation method of the paraffin section of pear flower tissue, which comprises the steps of:
(1) pear flower saved in 4-5 degrees Celsius of refrigerator is taken out, it is clean with distilled water flushing, by pistil, stamen, calyx, flower
3mm below valve and anthocaulus base portion is all removed, and obtains treated pear flower tissue；
(2) the pear flower tissue obtained in step (1) is put into the FAA fixer of fixed container, fixed container is evacuated,
Until no longer there is bubble in FAA fixer, it is 1-2 days fixed after pumping；
(3) the pear flower tissue obtained after pumping is placed on gauze and blots FAA fixer, and be successively dehydrated, be transparent, leaching
Wax, embedding, slice, bonding die, dewaxing, dyeing and removal floating color processing.
2. preparation method according to claim 1, which is characterized in that in the step (3) the step of dehydration are as follows: will
Pear flower group after blotting FAA fixer is woven in cryopreservation tube each step by step according to 70%, 80%, 85%, 90%, 95% concentration of alcohol
Dehydration is primary, each 2h, then uses 95% dehydration of alcohol 1h, is repeated twice, and last dehydrated alcohol dehydration is twice, each every time
45min。
3. preparation method according to claim 1, which is characterized in that transparent step in the step (3) are as follows: be dehydrated
Cheng Hou reprocesses twice sample according to the sequence of 1/2 countless+1/2 dimethylbenzene of ethyl alcohol, pure dimethylbenzene, transparent every time
Time is 1.5h.
4. preparation method according to claim 1, which is characterized in that the step of waxdip is handled in the step (3) are as follows:
Oven temperature is adjusted to 80 degrees Celsius, is put into paraffin melting spare；
Prepare cryopreservation tube, after 1/2 dimethylbenzene is added, is put into baking oven, the paraffin of thawing is added in cryopreservation tube after 30 seconds, will freeze
It deposits pipe lid to seal, be placed in 60 degrees Celsius of baking ovens；
From baking oven take out cryopreservation tube open, be put into complete it is transparent after material, sealing, when continuing to be put into 60 degrees Celsius of baking ovens
Between be 16h；
Material taking-up is put into pure wax liquor, at 60 c, every 2h is changed once, and replacement is twice.
5. preparation method according to claim 1, which is characterized in that in the step (3) the step of embedding treatment are as follows:
Imbedded mold is folded using brown paper；
Water-bath is set as 80 degrees Celsius, and beaker is placed in water-bath and is preheated, then in beaker be added 80% pure wax with
And 20% old wax；
One block of cardboard is placed as station on water-bath top, and embedded box is placed on cardboard, half paraffin is poured into, by material
It takes out, is put into embedded box from baking oven；
Material angle is adjusted using tweezers, with outside at calyx, apart from paper wall 1cm, two materials are at a distance of 2cm；It is fallen after adjustment
Enter other half paraffin, embedded box is placed on cooling, is cooled to upper layer and occurs continuously adding paraffin extremely when a tunic
Highest point.
6. preparation method according to claim 1, which is characterized in that in the step (3) the step of slice are as follows:
Embedded box is opened, is trimmed to paraffin mass using pocket knife trapezoidal, is bottom at calyx, is upper bottom at anthocaulus, when slice
Firmly uniformly, notch is vertical, and small trapezoidal each face is smooth, and 1cm paraffin is reserved at anthocaulus, and paraffin is as few as possible at calyx, remaining is each
Place reserves 2mm paraffin；
The paraffin reserved at anthocaulus is sticked on wood particle, guarantees that entire material is vertical with adhesive surface, is then fixed on manual pulley
On rotatable slicer, setting slice thickness is 10 microns, keeps vertical with blade at calyx；
In slicing processes, since pear flower tissue is larger, and each position size is uneven, and the dynamics for shaking slice handle is appropriate, from upper
Portion turns down rapidly to when incision, from bottom to top when slow but speed it is uniform.
7. preparation method according to claim 1, which is characterized in that bonding die step in the step (3) are as follows:
Use distilled water flushing glass slide, baking oven drying；Bonding die agent is obtained by filtration in egg white, is placed in 4 degrees Celsius of refrigerators and protects
It deposits；
Bonding die agent is applied on glass slide, while adding 6-8 drop distilled water, it is applied using glass bar, guarantee have on glass slide
Bonding die agent is placed in bonding die agent downward by the reverse side of paraffin section at this time；
After placing, toasted using alcolhol burner in glass slide lower part, the paraffin around discovery slice disappears, and is sure not to heat
Excessively, so that tissue is destroyed；
Distilled water is sucked using filter paper after baking piece, group is woven in open and flat on glass slide at this time, and paraffin is covered on tissue surface, and organizes
It is tightly combined with glass slide.
8. preparation method according to claim 7, which is characterized in that in the step (3) the step of dewaxing are as follows: glass will be carried
Piece is put into pure dimethylbenzene, and dewax 4h, 3h, 2h；After guaranteeing that dewaxing effect, dimethylbenzene use 4-5 times, need replacing.
9. preparation method according to claim 7, which is characterized in that in the step (3) the step of dyeing are as follows:
After the completion of dewaxing, will slice is repeated twice according to+1/2 dehydrated alcohol of 1/2 paraffin, without watery wine, 95% alcohol is repeated twice,
The sequence progress rehydration processing of 90% alcohol, 85% alcohol, 80% alcohol, 70% alcohol, 50% alcohol, 3 minutes every time；
Glass slide after rehydration is placed on 1% sarranine solution dyeing 16h, glass slide is taken out and is placed in 85% alcohol and impregnate 5 seconds,
0.5% fast green solution is subsequently placed into dye 5 seconds.
10. preparation method according to claim 7, which is characterized in that in the step (3) the step of removal floating color are as follows: dye
After the completion of color, glass slide is put in 90% alcohol, 95% alcohol is repeated twice, dehydrated alcohol is repeated twice middle removal floating color, every time
15 seconds；By glass slide be put into pure dimethylbenzene it is transparent after ten minutes, it is micro- pear flower tissue can be observed under an optical microscope
Structure can clearly be found, be dyed to the bast of red xylem and green.
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| CN109991063A (en) * | 2019-04-25 | 2019-07-09 | 广西壮族自治区药用植物园 | The paraffin section method of Paris polyphylla root tuber |
| CN111141568A (en) * | 2019-12-31 | 2020-05-12 | 华南农业大学 | Method for obtaining phloem of fast-growing tree species and application thereof |
| CN111257089A (en) * | 2020-02-18 | 2020-06-09 | 广东省农业科学院环境园艺研究所 | Optimized manufacturing method of paraffin sections of different flower tissues of butterfly orchid |
| CN112284791A (en) * | 2020-10-16 | 2021-01-29 | 扬州大学 | Method for separating xylem duct of grape fruit |
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