CN109187127A - Observe the preparation method of the slice dyed blended liquid and slice of xylem parenchyma - Google Patents
Observe the preparation method of the slice dyed blended liquid and slice of xylem parenchyma Download PDFInfo
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- CN109187127A CN109187127A CN201811050000.7A CN201811050000A CN109187127A CN 109187127 A CN109187127 A CN 109187127A CN 201811050000 A CN201811050000 A CN 201811050000A CN 109187127 A CN109187127 A CN 109187127A
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- 239000007788 liquid Substances 0.000 title claims abstract description 62
- 238000002360 preparation method Methods 0.000 title claims abstract description 30
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 220
- 238000004043 dyeing Methods 0.000 claims abstract description 57
- 239000000243 solution Substances 0.000 claims abstract description 51
- 239000007864 aqueous solution Substances 0.000 claims abstract description 40
- 241001062009 Indigofera Species 0.000 claims abstract description 39
- 235000019441 ethanol Nutrition 0.000 claims description 81
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 35
- 230000018044 dehydration Effects 0.000 claims description 21
- 238000006297 dehydration reaction Methods 0.000 claims description 21
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 claims description 16
- 229910002092 carbon dioxide Inorganic materials 0.000 claims description 8
- 239000001569 carbon dioxide Substances 0.000 claims description 8
- 239000007787 solid Substances 0.000 claims description 8
- 244000283070 Abies balsamea Species 0.000 claims description 6
- 235000007173 Abies balsamea Nutrition 0.000 claims description 6
- 239000004858 Canada balsam Substances 0.000 claims description 6
- 239000011521 glass Substances 0.000 claims description 6
- 238000007654 immersion Methods 0.000 claims description 5
- 239000008213 purified water Substances 0.000 claims description 5
- 239000002904 solvent Substances 0.000 claims description 4
- 239000008367 deionised water Substances 0.000 claims description 2
- 229910021641 deionized water Inorganic materials 0.000 claims description 2
- 239000012153 distilled water Substances 0.000 claims description 2
- 230000000694 effects Effects 0.000 abstract description 24
- 238000000034 method Methods 0.000 abstract description 17
- 239000000975 dye Substances 0.000 abstract description 12
- 239000003086 colorant Substances 0.000 abstract description 4
- 239000000835 fiber Substances 0.000 abstract description 2
- 239000012174 chinese wax Substances 0.000 description 24
- 244000044283 Toxicodendron succedaneum Species 0.000 description 14
- 238000004040 coloring Methods 0.000 description 9
- 241000530115 Clerodendrum trichotomum Species 0.000 description 8
- 235000011511 Diospyros Nutrition 0.000 description 8
- 244000236655 Diospyros kaki Species 0.000 description 8
- 241000931143 Gleditsia sinensis Species 0.000 description 8
- 241000894007 species Species 0.000 description 6
- 229950003937 tolonium Drugs 0.000 description 6
- 239000000463 material Substances 0.000 description 5
- 238000004140 cleaning Methods 0.000 description 4
- 230000000052 comparative effect Effects 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 238000000879 optical micrograph Methods 0.000 description 4
- HNONEKILPDHFOL-UHFFFAOYSA-M tolonium chloride Chemical compound [Cl-].C1=C(C)C(N)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 HNONEKILPDHFOL-UHFFFAOYSA-M 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 3
- 230000004069 differentiation Effects 0.000 description 3
- OARRHUQTFTUEOS-UHFFFAOYSA-N safranin Chemical compound [Cl-].C=12C=C(N)C(C)=CC2=NC2=CC(C)=C(N)C=C2[N+]=1C1=CC=CC=C1 OARRHUQTFTUEOS-UHFFFAOYSA-N 0.000 description 3
- RWVGQQGBQSJDQV-UHFFFAOYSA-M sodium;3-[[4-[(e)-[4-(4-ethoxyanilino)phenyl]-[4-[ethyl-[(3-sulfonatophenyl)methyl]azaniumylidene]-2-methylcyclohexa-2,5-dien-1-ylidene]methyl]-n-ethyl-3-methylanilino]methyl]benzenesulfonate Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C(=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C)C=2C(=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C)C=C1 RWVGQQGBQSJDQV-UHFFFAOYSA-M 0.000 description 3
- 238000010186 staining Methods 0.000 description 3
- RZSYLLSAWYUBPE-UHFFFAOYSA-L Fast green FCF Chemical compound [Na+].[Na+].C=1C=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C(=CC(O)=CC=2)S([O-])(=O)=O)C=CC=1N(CC)CC1=CC=CC(S([O-])(=O)=O)=C1 RZSYLLSAWYUBPE-UHFFFAOYSA-L 0.000 description 2
- ZXJXZNDDNMQXFV-UHFFFAOYSA-M crystal violet Chemical compound [Cl-].C1=CC(N(C)C)=CC=C1[C+](C=1C=CC(=CC=1)N(C)C)C1=CC=C(N(C)C)C=C1 ZXJXZNDDNMQXFV-UHFFFAOYSA-M 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 229960001235 gentian violet Drugs 0.000 description 2
- RNVCVTLRINQCPJ-UHFFFAOYSA-N o-toluidine Chemical compound CC1=CC=CC=C1N RNVCVTLRINQCPJ-UHFFFAOYSA-N 0.000 description 2
- QCDYQQDYXPDABM-UHFFFAOYSA-N phloroglucinol Chemical compound OC1=CC(O)=CC(O)=C1 QCDYQQDYXPDABM-UHFFFAOYSA-N 0.000 description 2
- 229960001553 phloroglucinol Drugs 0.000 description 2
- 238000009971 piece dyeing Methods 0.000 description 2
- 239000002023 wood Substances 0.000 description 2
- JPYHHZQJCSQRJY-UHFFFAOYSA-N Phloroglucinol Natural products CCC=CCC=CCC=CCC=CCCCCC(=O)C1=C(O)C=C(O)C=C1O JPYHHZQJCSQRJY-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 230000002380 cytological effect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- WPPDXAHGCGPUPK-UHFFFAOYSA-N red 2 Chemical compound C1=CC=CC=C1C(C1=CC=CC=C11)=C(C=2C=3C4=CC=C5C6=CC=C7C8=C(C=9C=CC=CC=9)C9=CC=CC=C9C(C=9C=CC=CC=9)=C8C8=CC=C(C6=C87)C(C=35)=CC=2)C4=C1C1=CC=CC=C1 WPPDXAHGCGPUPK-UHFFFAOYSA-N 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 238000010025 steaming Methods 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
Classifications
-
- 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
- G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
-
- 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
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
-
- 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
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
- G01N2001/302—Stain compositions
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Pathology (AREA)
- General Health & Medical Sciences (AREA)
- Immunology (AREA)
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Physics & Mathematics (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Cosmetics (AREA)
- Coloring (AREA)
- Sampling And Sample Adjustment (AREA)
Abstract
The invention discloses the preparation methods of the slice dyed blended liquid and slice of a kind of observation xylem parenchyma.The dyed blended liquid of the slice includes: sarranine ethanol solution and A Erxin indigo plant aqueous solution；Wherein, the volume ratio of sarranine ethanol solution and A Erxin indigo plant aqueous solution is 30~40:60~70.Wherein, the concentration of the sarranine ethanol solution and A Erxin indigo plant aqueous solution is 0.008~0.012g/ml.Sarranine coloured fibre tissue in the dyed blended liquid of the slice, A Erxin indigo plant dyes parenchymal tissue, parenchymal tissue, fibr tissue and the conduit (or tracheid) of slice after dyeing can be clearly observed simultaneously, xvlem fibres tissue and parenchymal tissue can obviously be distinguished, dyeing effect compared to other sarranine ethanol solutions and the dyeing liquor of A Erxin indigo plant aqueous solution volume ratio is more preferable, and slice preparation time is short, suitable for all xylophytas, overcomes coloring agent in traditional dicing method and be difficult to the shortcomings that distinguishing both tissues by color.
Description
Technical field
The present invention relates to a kind of technical fields of biologic slice preparation, more particularly to a kind of observation xylem parenchyma group
The preparation method of the slice dyed blended liquid and slice knitted.
Background technique
Production xylem slice is the conventional means of phytotomy research.The xylem of xylophyta is mainly by conduit
(or tracheid), fibr tissue and parenchymal tissue composition.Parenchymal tissue is the living cells in xylem, have storage, transmitting water and
The function of nutriment, in order to reinforce the research to plant inter-species function, tactful difference, there is an urgent need to thin to xylophyta now
The character of wall tissue is studied.
But the coloring agent of currently used production xylem slice is all pure to the coloring of parenchymal tissue and fibr tissue
The differentiation of the color of color or the two is unobvious, so being difficult to distinguish parenchymal tissue and fibr tissue by color.Such as it is common
Coloring agent has phloroglucin (phloroglucinol), gentian violet (gentian violet), toluidine blue (toluidine
Blue) and the mixed liquor comprising sarranine (safranin) and fast green (fast green) all can not highly significant, intuitively distinguish
Open fibr tissue and parenchymal tissue.
Prepare paraffin section, the method for resin slicer require be added embedding medium, the dicing method of embedding medium is added often
The processes such as transparent, embedding, slice, dewaxing (de- embedding medium), dyeing, mounting are needed, preparation process is complex, often a lot sample
Product, which are done, gets off to need to spend several days time, time-consuming very long.In addition, embedding medium immerse plant tissue can also destroy parenchymal tissue this
The activity of kind living cells, is further difficult to differentiate between parenchymal tissue and fibr tissue.
Sarranine, also referred to as safranin O, substantially red 2 or safranine T, are used in histology and cytological biological stain.A Er
Xin Lan, English name: Alcian Blue, CAS:12040-44-7: molecular formula: C56H68Cl4CuN16S4, Chinese nickname: A Erxin
Blue, alcian blue, has the following structure formula:
Summary of the invention
The purpose of the present invention is to provide a kind of dyed blended liquid of slice for observing xylem parenchyma, are able to solve
Not the problem of existing xylem microsection manufacture method complexity, the long and thin wall tissue of short time consumption and fibr tissue are not easily distinguishable.
To achieve the above object, the present invention provides a kind of dyed blended liquid of slice for observing xylem parenchyma, packets
It includes: sarranine ethanol solution and A Erxin indigo plant aqueous solution；Wherein, the volume ratio of sarranine ethanol solution and A Erxin indigo plant aqueous solution is 30
~40:60~70.
In a preferred embodiment, the concentration of the sarranine ethanol solution is 0.008~0.012g/ml, A Erxin
The concentration of blue aqueous solution is 0.008~0.012g/ml；Preferably, the concentration of the sarranine ethanol solution be 0.009~
The concentration of 0.011g/ml, A Erxin indigo plant aqueous solution is 0.009~0.011g/ml；Most preferably, the sarranine ethanol solution
Concentration is 0.010g/ml, and the concentration of A Erxin indigo plant aqueous solution is 0.010g/ml.
In a preferred embodiment, the solvent of the sarranine ethanol solution be concentration of volume percent be 40%~
60% ethanol solution；Preferably, the concentration of volume percent of the ethanol solution is 50%.
Sarranine solute effect can be made good using the concentration ethanol solution, solution rate is fast, and dissolution is thoroughly and without residue.
In a preferred embodiment, the solvent of the A Erxin indigo plant aqueous solution is selected from pure water, deionized water or steaming
Any one in distilled water.
In a preferred embodiment, the volume ratio of the sarranine ethanol solution and A Erxin indigo plant aqueous solution be 32~
37:62~67；Preferably, the volume ratio of the sarranine ethanol solution and A Erxin indigo plant aqueous solution is 35:65.
Another object of the present invention is to provide a kind of preparation method for being sliced dyed blended liquid, the slice is dyed blended
Liquid is that any one of the above is sliced dyed blended liquid, and preparation step includes: to dissolve sarranine using ethyl alcohol, and being configured to concentration is
The sarranine ethanol solution of 0.008~0.012g/ml；It is blue using dissolved in purified water A Erxin, be configured to concentration be 0.008~
The A Erxin indigo plant aqueous solution of 0.012g/ml；By the sarranine ethanol solution and A Erxin indigo plant aqueous solution 30~40:60 by volume
~70 ratio is mixed, and the dyed blended liquid of slice is configured to.
Another object of the present invention is to provide a kind of preparation method of slice, the slice is xylem slice, including
Following steps:
(1) fresh xylem sample is impregnated into water suction in water, the xylem sample after being absorbed water；
(2) the xylem sample after impregnating step (1) cuts out cross-sectional slices；And
(3) it gained slice is immersed into any one of the above being sliced in dyed blended liquid and dye, dyeing time is 3~
Then 5min is dehydrated with graded ethanol solutions, the xylem slice after obtaining dehydration dyeing.
In a preferred embodiment, in above-mentioned steps (1), the time that xylem sample impregnates in water be 3min~
6h。
In a preferred embodiment, in above-mentioned steps (2), the thickness of the cross-sectional slices is 18~20 μm.
In a preferred embodiment, in above-mentioned steps (3), the concentration of volume percent of the graded ethanol solutions is
1~100%.
In a preferred embodiment, in above-mentioned steps (3), the slice after dyeing is first cleaned with pure water, so
Solid carbon dioxide point is dipped on blotting paper afterwards, then is dehydrated respectively with 50% ethyl alcohol, 75% ethyl alcohol, 95% ethyl alcohol, is taken out de-
Slice after water dips in solid carbon dioxide point and ethyl alcohol on blotting paper；Preferably, the dewatering time of each graded ethanol is 3~5s.
In a preferred embodiment, further the xylem slice after dehydration dyeing obtained by above-mentioned steps (3) is put
It is placed on glass slide, canada balsam is added dropwise, covered flattens, permanent section is made.
Compared with prior art, the invention has the following beneficial effects:
(1) present invention is sliced sarranine coloured fibre tissue in dyed blended liquid, and A Erxin indigo plant dyes parenchymal tissue, the mixing
Parenchymal tissue, the fibr tissue of wooden cross section can be clearly observed in xylem slice after dyeing liquor dyeing simultaneously
With conduit (or tracheid), the fibr tissue and parenchymal tissue of xylem can be significantly distinguished, is overcome in traditional dicing method
Coloring agent is difficult to the shortcomings that both tissues are distinguished by color.
(2) present invention is the sarranine ethanol solution of 30~40:60~70 and mixing for A Erxin indigo plant aqueous solution using volume ratio
It closes dyeing liquor to dye xylem slice, slice imaging effect dyes far better than no dyeing liquid and only passes through single dye
The effect of color liquid dyeing, the parenchymal tissue and fibr tissue of the xylem slice of no dyeing liquid dyeing are only led to without colouring discrimination
The parenchymal tissue and fibr tissue for crossing the xylem slice of single dyeing liquor dyeing are a kind of color, and the two can not pass through differentiation
Parenchymal tissue and fibr tissue；In addition, using the volume ratio sarranine ethanol solution and A Erxin indigo plant aqueous solution it is dyed blended
Liquid phase is more preferable to the dyeing effect of the dyed blended liquid of other sarranine ethanol solutions and A Erxin indigo plant aqueous solution volume ratio, using it
The thin-walled group of dyeing xylem slice obtained by the sarranine ethanol solution of his volume ratio and the dyed blended liquid of A Erxin indigo plant aqueous solution
It knits unobvious with the colouring discrimination of fibr tissue, cannot clearly distinguish the fibr tissue and parenchymal tissue of xylem, and use body
Product is than being the thin-walled group being sliced obtained by the sarranine ethanol solution of 30~40:60~70 and the dyed blended liquid of A Erxin indigo plant aqueous solution
It knits with the colouring discrimination of fibr tissue clearly.
(3) compared to other existing frequently-used slice preparation methods, this method is not required to embedded material, can directly quick Fabrication at
Permanent section is used for later observation, and entire preparation time is no more than 10 minutes, and the entire preparation time that is sliced is short, is suitable for all wood
This plant can be applied in the anatomical research of xylophyta.
Detailed description of the invention
Fig. 1 is the Chinese wax tree species xylem that microscopically observation obtains after the dyeing of the dyed blended liquid of the embodiment of the present invention 1
The effects of cross-sectional slices shows figure, and wherein parenchymal tissue is blue, fibr tissue takes on a red color；
Fig. 2 is the clerodendron trichotomum tree species wood that microscopically observation obtains after the dyeing of the dyed blended liquid of the embodiment of the present invention 1
The effect of matter cross section slice shows figure, and wherein parenchymal tissue is blue, fibr tissue takes on a red color；
Fig. 3 is the persimmon tree species xylem that microscopically observation obtains after the dyeing of the dyed blended liquid of the embodiment of the present invention 1
The effects of cross-sectional slices shows figure, and wherein parenchymal tissue is blue, fibr tissue takes on a red color；
Fig. 4 is the Chinese honey locust tree species xylem that microscopically observation obtains after the dyeing of the dyed blended liquid of the embodiment of the present invention 1
The effects of cross-sectional slices shows figure, and wherein parenchymal tissue is blue, fibr tissue takes on a red color；
Fig. 5 is the Chinese wax tree species xylem cross-sectional slices that example 1 is obtained without microscopically observation after dyeing by contrast
Effect display figure；
Fig. 6 is that the Chinese wax tree species xylem that microscopically observation obtains after 2 Toluidine blue staining liquid of example dyeing by contrast is horizontal
The effect of cross-sectional slice shows figure；
Fig. 7 is the obtained Chinese wax tree species xylem cross section of microscopically observation after the dyed blended liquid A of example 3 dyeing by contrast
The effect of slice shows figure；
Fig. 8 is the obtained Chinese wax tree species xylem cross section of microscopically observation after the dyed blended liquid B of example 4 dyeing by contrast
The effect of slice shows figure.
Main appended drawing reference explanation:
1- parenchymal tissue, 2- fibr tissue, 3- conduit.
Specific embodiment
With reference to the accompanying drawing, specific embodiments of the present invention will be described in detail, it is to be understood that guarantor of the invention
Shield range is not limited by the specific implementation.
Unless otherwise explicitly stated, otherwise in entire disclosure and claims, term " includes " or its change
Changing such as "comprising" or " including " etc. will be understood to comprise stated element or component, and not exclude other members
Part or other component parts.
Unless otherwise specified, various raw material, reagent, the instrument and equipment etc. used in the present invention can pass through city
Field is commercially available or can be prepared by existing method.
Sarranine: it is purchased from Suo Laibao (Solarbio), article number: S8020；
A Erxin indigo plant (Alcian Blue): it is purchased from Suo Laibao (Solarbio), article number: A8140.
A kind of embodiment 1: preparation method of the dyed blended liquid for xylem slice dyeing
Preparation step includes: that concentration of volume percent is used to dissolve sarranine for 50% ethyl alcohol, and being configured to concentration is
The sarranine ethanol solution of 0.010g/ml；It is blue using dissolved in purified water A Erxin, it is configured to the A Erxin that concentration is 0.010g/ml
Blue aqueous solution；By 35: 65 mixing by volume of sarranine ethanol solution and A Erxin indigo plant aqueous solution, it is configured to dyed blended liquid.
A kind of embodiment 2: preparation method of the dyed blended liquid for xylem slice dyeing
Preparation step includes: that concentration of volume percent is used to dissolve sarranine for 50% ethyl alcohol, and being configured to concentration is
The sarranine ethanol solution of 0.010g/ml；It is blue using dissolved in purified water A Erxin, it is configured to the A Erxin that concentration is 0.010g/ml
Blue aqueous solution；By 40: 60 mixing by volume of sarranine ethanol solution and A Erxin indigo plant aqueous solution, it is configured to dyed blended liquid.
A kind of embodiment 3: preparation method of the dyed blended liquid for xylem slice dyeing
Preparation step includes: that concentration of volume percent is used to dissolve sarranine for 50% ethyl alcohol, and being configured to concentration is
The sarranine ethanol solution of 0.010g/ml；It is blue using dissolved in purified water A Erxin, it is configured to the A Erxin that concentration is 0.010g/ml
Blue aqueous solution；By 30: 70 mixing by volume of sarranine ethanol solution and A Erxin indigo plant aqueous solution, it is configured to dyed blended liquid.
A kind of embodiment 4: colouring method of Chinese wax xylem slice rapid dyeing
Preparation step includes: that Chinese wax xylem sample is impregnated 30min/h in water, so that specimen material sufficiently absorbs water；
Chinese wax xylem sample after immersion is cut out into the cross-sectional slices with a thickness of 18 μm；Take 1 gained of 1ml embodiment dyed blended
Gained slice is immersed and dyes by liquid, dyeing time 3min；Then it is dipped in after cleaning 3s with pure water with blotting paper dry
The moisture of slice；Ethyl alcohol that the ethyl alcohol for being respectively again 50% with concentration of volume percent, concentration of volume percent are 75%, volume
The ethanol dehydration that percent concentration is 95%, each ethanol dehydration time is 3s, dips in solid carbon dioxide point and second after taking-up on blotting paper
Alcohol, the Chinese wax xylem slice after obtaining dehydration dyeing.
Chinese wax xylem slice after gained dehydration dyeing is paved on glass slide, canada balsam is added dropwise, closes the lid
Slide flattens, permanent section is made, and in optical microphotograph microscopic observation, takes pictures, and obtains Fig. 1 and dyes through the dyed blended liquid of embodiment 1
The effect display figure for the Chinese wax tree species xylem cross-sectional slices that microscopically observation obtains afterwards.It was found from the figure that parenchymal tissue
Blue in a slice, fibr tissue takes on a red color in a slice, can be clearly observed very much the thin-walled of wooden cross section
Tissue, fibr tissue and conduit, can obviously distinguish the fibr tissue and parenchymal tissue of Chinese wax xylem.
A kind of embodiment 5: colouring method of clerodendron trichotomum xylem slice rapid dyeing
Preparation step includes: that clerodendron trichotomum xylem sample is impregnated 30min/h in water, so that specimen material is sufficiently inhaled
Water；Clerodendron trichotomum xylem sample after immersion is cut out into the cross-sectional slices with a thickness of 18 μm；It takes and is mixed obtained by 1ml embodiment 1
Dyeing liquor is closed, gained slice is immersed and is dyed, dyeing time 3min；Then with water suction after cleaning 3s with pure water
Paper dips in the moisture of dry chip；The second that the ethyl alcohol for being respectively again 50% with concentration of volume percent, concentration of volume percent are 75%
Alcohol, the ethanol dehydration that concentration of volume percent is 95%, each ethanol dehydration time is 3s, dips in solid carbon dioxide after taking-up on blotting paper
Divide and ethyl alcohol, the clerodendron trichotomum xylem after obtaining dehydration dyeing are sliced.
Clerodendron trichotomum xylem slice after gained dehydration dyeing is paved on glass slide, canada balsam is added dropwise, covers
Upper coverslip flattens, permanent section is made, and in optical microphotograph microscopic observation, takes pictures, obtains Fig. 2 through the dyed blended liquid of embodiment 1
The effect display figure for the clerodendron trichotomum tree species xylem cross-sectional slices that microscopically observation obtains after dyeing.It was found from the figure that
Parenchymal tissue is blue in a slice, and fibr tissue takes on a red color in a slice, and it is transversal can be very clearly observed xylem
Parenchymal tissue, fibr tissue and the conduit in face can obviously distinguish the fibr tissue and parenchymal tissue of clerodendron trichotomum xylem.
A kind of embodiment 6: colouring method of persimmon xylem slice rapid dyeing
Preparation step includes: that persimmon xylem sample is impregnated 30min/h in water, so that specimen material sufficiently absorbs water；
Persimmon xylem sample after immersion is cut out into the cross-sectional slices with a thickness of 18 μm；Take 1 gained of 1ml embodiment dyed blended
Gained slice is immersed and dyes by liquid, dyeing time 3min；Then it is dipped in after cleaning 3s with pure water with blotting paper dry
The moisture of slice；Ethyl alcohol that the ethyl alcohol for being respectively again 50% with concentration of volume percent, concentration of volume percent are 75%, volume
The ethanol dehydration that percent concentration is 95%, each ethanol dehydration time is 3s, dips in solid carbon dioxide point and second after taking-up on blotting paper
Alcohol, the persimmon xylem slice after obtaining dehydration dyeing.
Persimmon xylem slice after gained dehydration dyeing is paved on glass slide, canada balsam is added dropwise, closes the lid
Slide flattens, permanent section is made, and in optical microphotograph microscopic observation, takes pictures, and obtains Fig. 3 and dyes through the dyed blended liquid of embodiment 1
The effect display figure for the persimmon tree species xylem cross-sectional slices that microscopically observation obtains afterwards.It was found from the figure that parenchymal tissue
Blue in a slice, fibr tissue takes on a red color in a slice, can be clearly observed very much the thin-walled of wooden cross section
Tissue, fibr tissue and conduit, can obviously distinguish the fibr tissue and parenchymal tissue of persimmon xylem.
A kind of embodiment 7: colouring method of Chinese honey locust xylem slice rapid dyeing
Preparation step includes: that Chinese honey locust xylem sample is impregnated 30min/h in water, so that specimen material sufficiently absorbs water；
Chinese honey locust xylem sample after immersion is cut out into the cross-sectional slices with a thickness of 18 μm；Take 1 gained of 1ml embodiment dyed blended
Gained slice is immersed and dyes by liquid, dyeing time 3min；Then it is dipped in after cleaning 3s with pure water with blotting paper dry
The moisture of slice；Ethyl alcohol that the ethyl alcohol for being respectively again 50% with concentration of volume percent, concentration of volume percent are 75%, volume
The ethanol dehydration that percent concentration is 95%, each ethanol dehydration time is 3s, dips in solid carbon dioxide point and second after taking-up on blotting paper
Alcohol, the Chinese honey locust xylem slice after obtaining dehydration dyeing.
Chinese honey locust xylem slice after gained dehydration dyeing is paved on glass slide, canada balsam is added dropwise, closes the lid
Slide flattens, permanent section is made, and in optical microphotograph microscopic observation, takes pictures, and obtains Fig. 4 and dyes through the dyed blended liquid of embodiment 1
The effect display figure for the Chinese honey locust tree species xylem cross-sectional slices that microscopically observation obtains afterwards.It was found from the figure that parenchymal tissue
Blue in a slice, fibr tissue takes on a red color in a slice, can be clearly observed very much the thin-walled of wooden cross section
Tissue, fibr tissue and conduit, can obviously distinguish the fibr tissue and parenchymal tissue of Chinese honey locust xylem.
Comparative example 1: slice is not dyed
Directly 4 identical method processing Chinese wax xylems are sliced through the foregoing embodiment, but are not dyed to slice,
The effect for obtaining Fig. 5 Chinese wax tree species xylem cross-sectional slices that example 1 is obtained without microscopically observation after dyeing by contrast is aobvious
Diagram.
It was found from the figure that the xylem parenchyma and fibr tissue of Chinese wax tree species be without colouring discrimination, to cannot be distinguished.
Comparative example 2: slice is dyed using Toluidine blue staining liquid common in anatomical research
4 identical methods processing Chinese wax xylems slice through the foregoing embodiment, except that the dyeing liquor used for
Toluidine blue obtains Fig. 6 obtained Chinese wax tree species xylem of microscopically observation after the dyeing of 2 Toluidine blue staining liquid of example by contrast
The effect of cross-sectional slices shows figure.
It was found from the figure that the xylem parenchyma and fibr tissue of Chinese wax tree species are blue, color area can not be passed through
Divide parenchymal tissue and fibr tissue.
Comparative example 3: using the sarranine ethanol solution of different volumes ratio and the dyed blended liquid of A Erxin indigo plant aqueous solution to cutting
Piece dyeing
4 identical methods processing Chinese wax xylems slice through the foregoing embodiment, except that the dyeing liquor used for
Dyed blended liquid A obtains Fig. 7 obtained Chinese wax tree species xylem of microscopically observation after the dyed blended liquid A of example 3 dyeing by contrast
The effect of cross-sectional slices shows figure.Wherein, dyed blended liquid A includes: the sarranine ethanol solution and 0.010g/ of 0.010g/ml
The volume ratio of the A Erxin indigo plant aqueous solution of ml, sarranine ethanol solution and A Erxin indigo plant aqueous solution is 15:95.
It was found from the figure that the xylem parenchyma and fibr tissue color of Chinese wax tree species cannot be distinguished without significant difference
Both tissues.
Comparative example 4: using the sarranine ethanol solution of different volumes ratio and the dyed blended liquid of A Erxin indigo plant aqueous solution to cutting
Piece dyeing
Using above-mentioned dyed blended liquid 4, and 4 identical method processing Chinese wax xylem slice through the foregoing embodiment, it obtains
The effect for the Chinese wax tree species xylem cross-sectional slices that microscopically observation obtains after to Fig. 8 by contrast dyed blended liquid B of example 4 dyeing
Fruit display figure.Wherein, the dyed blended liquid B of use includes: the sarranine ethanol solution of 0.010g/ml and the A Er of 0.010g/ml
The volume ratio of pungent indigo plant aqueous solution, sarranine ethanol solution and A Erxin indigo plant aqueous solution is 50:50.
It was found from the figure that the xylem parenchyma and fibr tissue of Chinese wax tree species can be distinguished by color reluctantly, still
Effect is clear not as good as Fig. 1 that embodiment 4 obtains, and it is obvious that differentiation effect is also not so good as Fig. 1.
The aforementioned description to specific exemplary embodiment of the invention is in order to illustrate and illustration purpose.These descriptions
It is not wishing to limit the invention to disclosed precise forms, and it will be apparent that according to the above instruction, can much be changed
And variation.The purpose of selecting and describing the exemplary embodiment is that explaining specific principle of the invention and its actually answering
With so that those skilled in the art can be realized and utilize a variety of different exemplary implementation schemes of the invention and
Various chooses and changes.The scope of the present invention is intended to be limited by claims and its equivalents.
Claims (10)
1. a kind of dyed blended liquid of slice for observing xylem parenchyma, it is characterised in that: the dyed blended liquid packet of slice
It includes: sarranine ethanol solution and A Erxin indigo plant aqueous solution；Wherein, the volume ratio of sarranine ethanol solution and A Erxin indigo plant aqueous solution is 30
~40:60~70.
2. the dyed blended liquid of slice according to claim 1, it is characterised in that: the concentration of the sarranine ethanol solution is
The concentration of 0.008~0.012g/ml, A Erxin indigo plant aqueous solution is 0.008~0.012g/ml；Preferably, the sarranine ethyl alcohol is molten
The concentration of liquid is 0.009~0.011g/ml, and the concentration of A Erxin indigo plant aqueous solution is 0.009~0.011g/ml；Most preferably, institute
The concentration for stating sarranine ethanol solution is 0.010g/ml, and the concentration of A Erxin indigo plant aqueous solution is 0.010g/ml.
3. the dyed blended liquid of slice according to claim 1, it is characterised in that: the solvent of the sarranine ethanol solution is body
The ethanol solution that product percent concentration is 40%~60%；Preferably, the concentration of volume percent of the ethanol solution is 50%.
4. the dyed blended liquid of slice according to claim 1, it is characterised in that: the solvent of the A Erxin indigo plant aqueous solution selects
From in pure water, deionized water or distilled water any one.
5. the dyed blended liquid of slice according to claim 1, it is characterised in that: the sarranine ethanol solution and A Erxin are blue
The volume ratio of aqueous solution is 32~37:62~67；Preferably, the volume ratio of the sarranine ethanol solution and A Erxin indigo plant aqueous solution
It is 35:65.
6. a kind of preparation method for being sliced dyed blended liquid, the dyed blended liquid of slice is described in any one of Claims 1 to 5
Slice dyed blended liquid, preparation step includes: to dissolve sarranine using ethyl alcohol, and being configured to concentration is 0.008~0.012g/ml
Sarranine ethanol solution；It is blue using dissolved in purified water A Erxin, it is configured to the A Erxin indigo plant water that concentration is 0.008~0.012g/ml
Solution；The ratio of the sarranine ethanol solution and A Erxin indigo plant aqueous solution 30~40:60~70 by volume is mixed, is matched
The dyed blended liquid of slice is made.
7. a kind of preparation method of slice, the slice is that xylem is sliced, it is characterised in that: the preparation method includes following
Step:
(1) fresh xylem sample is impregnated into water suction in water, the xylem sample after being absorbed water；
(2) the xylem sample after impregnating step (1) cuts out cross-sectional slices；And
(3) gained is sliced to immerse in the dyed blended liquid of the described in any item slices of Claims 1 to 5 and is dyed, when dyeing
Between be 3~5min, be then dehydrated with graded ethanol solutions, obtain dehydration dyeing after xylem slice.
8. the preparation method of slice according to claim 7, it is characterised in that: in step (1), xylem sample is in water
The time of immersion is 3min~6h；
And/or in step (2), the thickness of the cross-sectional slices is 18~20 μm；
And/or in step (3), the concentration of volume percent of the graded ethanol solutions is 40~100%.
9. the preparation method of slice according to claim 7, it is characterised in that: in step (3), the slice after dyeing is first used
Pure water is cleaned, and solid carbon dioxide point is then dipped on blotting paper, then respectively with 50% ethyl alcohol, 75% ethyl alcohol, 95% second
Alcohol is dehydrated, and is taken out dewatered slice and is dipped in solid carbon dioxide point and ethyl alcohol on blotting paper；Preferably, the dehydration of each graded ethanol
Time is 3~5s.
10. the preparation method of slice according to claim 7, it is characterised in that: further contaminate dehydration obtained by step (3)
Xylem slice after color is placed on glass slide, and canada balsam is added dropwise, and covered flattens, permanent section is made.
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| CN201811050000.7A CN109187127A (en) | 2018-09-10 | 2018-09-10 | Observe the preparation method of the slice dyed blended liquid and slice of xylem parenchyma |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109822705A (en) * | 2019-03-11 | 2019-05-31 | 南京林业大学 | The colouring method of prothenchyma (of wood) and parenchyma cell in a kind of difference timber |
| CN110243568A (en) * | 2019-08-05 | 2019-09-17 | 中国空气动力研究与发展中心低速空气动力研究所 | A kind of low-speed wind tunnel sublimed method test method based on colored indicator |
| CN111060449A (en) * | 2019-12-30 | 2020-04-24 | 济宁明升新材料有限公司 | Novel method for judging discontinuous position of composite paper layer |
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- 2018-09-10 CN CN201811050000.7A patent/CN109187127A/en active Pending
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| D.TOLIVIA ET AL.: "a new polychromatic method for simultaneous and differential staining of plant tissues", 《JOURNAL OF MICROSCOPY》 * |
| YU ZHANG ET AL.: "Color Quantification of Stained Maize Stem Section Describes Lignin Spatial Distribution within the Whole Stem", 《AGRICULTURAL AND FOOD CHEMISTRY》 * |
| 程虎印: "《药用植物学实验指导》", 30 September 2014 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109822705A (en) * | 2019-03-11 | 2019-05-31 | 南京林业大学 | The colouring method of prothenchyma (of wood) and parenchyma cell in a kind of difference timber |
| CN110243568A (en) * | 2019-08-05 | 2019-09-17 | 中国空气动力研究与发展中心低速空气动力研究所 | A kind of low-speed wind tunnel sublimed method test method based on colored indicator |
| CN111060449A (en) * | 2019-12-30 | 2020-04-24 | 济宁明升新材料有限公司 | Novel method for judging discontinuous position of composite paper layer |
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