CN106501053B - The paraffin section method of meadowrueleaf corydalis root blade - Google Patents
The paraffin section method of meadowrueleaf corydalis root blade Download PDFInfo
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- CN106501053B CN106501053B CN201611262548.9A CN201611262548A CN106501053B CN 106501053 B CN106501053 B CN 106501053B CN 201611262548 A CN201611262548 A CN 201611262548A CN 106501053 B CN106501053 B CN 106501053B
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- 241000218176 Corydalis Species 0.000 title claims abstract description 58
- 238000000034 method Methods 0.000 title claims abstract description 46
- 239000012188 paraffin wax Substances 0.000 title claims abstract description 29
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims abstract description 69
- 230000001476 alcoholic effect Effects 0.000 claims abstract description 35
- 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 claims abstract description 31
- 238000004043 dyeing Methods 0.000 claims abstract description 25
- 238000000926 separation method Methods 0.000 claims abstract description 13
- 239000000975 dye Substances 0.000 claims abstract description 9
- 238000010186 staining Methods 0.000 claims abstract description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 242
- 230000018044 dehydration Effects 0.000 claims description 40
- 238000006297 dehydration reaction Methods 0.000 claims description 40
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 24
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 12
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 12
- 229920001661 Chitosan Polymers 0.000 claims description 11
- 239000002904 solvent Substances 0.000 claims description 11
- 239000003086 colorant Substances 0.000 claims description 4
- 238000012545 processing Methods 0.000 claims description 4
- 230000028514 leaf abscission Effects 0.000 claims description 3
- 239000002253 acid Substances 0.000 claims 1
- 150000004676 glycans Chemical class 0.000 claims 1
- 239000000052 vinegar Substances 0.000 claims 1
- 235000021419 vinegar Nutrition 0.000 claims 1
- 239000000463 material Substances 0.000 abstract description 27
- 239000001993 wax Substances 0.000 description 31
- 229960000583 acetic acid Drugs 0.000 description 21
- 239000011521 glass Substances 0.000 description 18
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 9
- 230000000694 effects Effects 0.000 description 9
- 241001411320 Eriogonum inflatum Species 0.000 description 6
- 238000007711 solidification Methods 0.000 description 6
- 230000008023 solidification Effects 0.000 description 6
- 239000003795 chemical substances by application Substances 0.000 description 4
- 239000007787 solid Substances 0.000 description 4
- 239000004575 stone Substances 0.000 description 4
- 235000002991 Coptis groenlandica Nutrition 0.000 description 3
- 244000247747 Coptis groenlandica Species 0.000 description 3
- 108010010803 Gelatin Proteins 0.000 description 3
- 210000001367 artery Anatomy 0.000 description 3
- 238000001816 cooling Methods 0.000 description 3
- 239000012153 distilled water Substances 0.000 description 3
- 239000000428 dust Substances 0.000 description 3
- 238000007667 floating Methods 0.000 description 3
- 229920000159 gelatin Polymers 0.000 description 3
- 239000008273 gelatin Substances 0.000 description 3
- 235000019322 gelatine Nutrition 0.000 description 3
- 235000011852 gelatine desserts Nutrition 0.000 description 3
- 239000012362 glacial acetic acid Substances 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 210000004932 little finger Anatomy 0.000 description 3
- 239000000155 melt Substances 0.000 description 3
- 229920001206 natural gum Polymers 0.000 description 3
- 238000005086 pumping Methods 0.000 description 3
- 238000006748 scratching Methods 0.000 description 3
- 230000002393 scratching effect Effects 0.000 description 3
- 239000008399 tap water Substances 0.000 description 3
- 235000020679 tap water Nutrition 0.000 description 3
- 238000010257 thawing Methods 0.000 description 3
- 210000004027 cell Anatomy 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 230000007774 longterm Effects 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000004321 preservation Methods 0.000 description 2
- 241000783421 Corydalis saxicola Species 0.000 description 1
- 241000876833 Emberizinae Species 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
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- 241000205578 Thalictrum Species 0.000 description 1
- 238000000862 absorption spectrum Methods 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000036592 analgesia Effects 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 230000007882 cirrhosis Effects 0.000 description 1
- 208000019425 cirrhosis of liver Diseases 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 208000006454 hepatitis Diseases 0.000 description 1
- 231100000283 hepatitis Toxicity 0.000 description 1
- 241000411851 herbal medicine Species 0.000 description 1
- 238000007654 immersion Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 239000010977 jade Substances 0.000 description 1
- 239000006028 limestone Substances 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000002574 poison Substances 0.000 description 1
- 231100000614 poison Toxicity 0.000 description 1
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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
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- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Sampling And Sample Adjustment (AREA)
Abstract
The invention discloses a kind of paraffin section methods of meadowrueleaf corydalis root blade, it fixes, be dehydrated including sample, is transparent, waxdip, embedding, slice and exhibition piece, dewaxing, dyeing and mounting, dewatered meadowrueleaf corydalis root blade is put in the sarranine solution that volumetric concentration is 0.8-1.2% and disseminates 8-12h by the dyeing the following steps are included: before transparent；In staining procedure, the slide for being loaded with meadowrueleaf corydalis root blade is subjected to rehydration, it is put in alcoholic solution again after rehydration and carries out color separation, the acetic acid that volume ratio is 1/ (800-1000) is added into the alcoholic solution for color separation simultaneously, is finally putting into the fast green solution that volumetric concentration is 0.8-1.2% and dyes 5-15 seconds.The meadowrueleaf corydalis root blade contour group that the present invention prepares knits the clear identification for being suitable for medicinal material.
Description
Technical field
The present invention relates to histology fields.It is more particularly related to a kind of paraffin section side of meadowrueleaf corydalis root blade
Method.
Background technique
Meadowrueleaf corydalis root (Corydalis saxicola Bunting) is bloodroot, is herbaceos perennial, distribution
It is saved in China Guangxi, Guangdong, Fujian, Hunan etc..All herbal medicine, herb contain deydrokaividing (Corydalis Saxicolae) isoreactivity ingredient,
It with significant antibacterial, anti-inflammatory, analgesia and stable effect by force, and finds that it plays the role of inhibiting tumour cell, it is swollen to cure mainly boil
The diseases such as poison, hepatitis, cirrhosis, liver cancer.Meadowrueleaf corydalis root is famous strong precious jade drug kind, and meadowrueleaf corydalis root plant distributions are confined to limestone
Area belongs to tor endemic species.Since habitat conditions is severe, natural propagation rate is very low, and population development is difficult, and resource reserves are very
Limited, in addition its seed is tiny, breeding is relatively difficult, and more difficult in introducing and planting to survive, people largely excavate and receive in recent years
Purchase, resulting in wild resource, supply falls short of demand, and wild meadowrueleaf corydalis root resource is on the verge of exhaustion.With the increasingly soaring market price, mix
The product that confuse emerge one after another, therefore are badly in need of developing the true and false of the meadowrueleaf corydalis root leaf sections to identify medicinal material, for specification medicinal material market, mirror
It is that meadowrueleaf corydalis root blade cross cut structure is clear that the meadowrueleaf corydalis root slice of the other true and false, which needs effect to be achieved, can clearly differentiate spongy tissue,
Palisade tissue, vein middle part, profile of vascular bundle etc., and need sample that can use with long-term preservation, it is permanent microscope slide
Sample.The many and diverse disunity of conventional section technical step, and the meadowrueleaf corydalis root blade contour group prepared is knitted and unintelligible can not be applicable in
In the identification of medicinal material.
Summary of the invention
It cannot it is an object of the invention to solve that meadowrueleaf corydalis root leaf tissue that conventional section technology prepares is unintelligible
To identify medicinal material authenticity, and provide the advantages of at least will be described later.
In order to realize these purposes and other advantages according to the present invention, a kind of paraffin section of meadowrueleaf corydalis root blade is provided
Method, including sample is fixed, is dehydrated, is transparent, waxdip, embedding, slice and exhibition piece, dewaxing, dyeing and mounting, the dyeing are wrapped
Include following steps:
Before transparent, dewatered meadowrueleaf corydalis root blade is put in the sarranine solution that volumetric concentration is 0.8-1.2% and is disseminated
8-12h；
In staining procedure, the slide that will be loaded with meadowrueleaf corydalis root blade carries out rehydration, be put in alcoholic solution again after rehydration into
Row color separation, while the acetic acid that volume ratio is 1/ (800-1000) being added into the alcoholic solution for color separation, it is finally putting into volume
It is dyed 5-15 seconds in the fast green solution that concentration is 0.8-1.2%.
Preferably, it is with+1/3 volume of 2/3 volume absolute alcohol that the volumetric concentration, which is the sarranine solution of 0.8-1.2%,
The mixed liquor of TO type biology clarifier is formulated for solvent, and the volumetric concentration is that the fast green solution of 0.8-1.2% is to use body
The alcoholic solution that product concentration is 95% is that solvent is formulated.
Preferably, before dyeing, the slide for being loaded with meadowrueleaf corydalis root blade after dewaxing is sequentially placed into the life of 1/2 volume type
+ 1/2 volume absolute alcohol mixed liquor of object clarifier, absolute alcohol, absolute alcohol, volumetric concentration are 95% alcohol, volumetric concentration
Be rehydration in 70% alcohol for 85% alcohol, volumetric concentration, every grade 3-5 seconds；It places into 50% alcohol color separation 3-10 seconds；Later
Be sequentially placed into that volumetric concentration is 70% alcohol, volumetric concentration is to be further processed in 85% alcohol again, every grade 3-10 seconds.
Preferably, the slide for being loaded with meadowrueleaf corydalis root blade is put in be put into absolute alcohol after fast green solution dyeing and washes away dye
Toner, this grade stop 3-10 seconds；Then absolute alcohol ,+1/2 absolute alcohol mixed liquor of 1/2TO type biology clarifier are sequentially placed into
In, every grade stop 3-10 seconds；Finally the slide for being loaded with meadowrueleaf corydalis root blade is put into TO type biology clarifier and is stopped 5-10 minutes,
This step is in triplicate.
Preferably, it will successively pass through that volumetric concentration is 80% alcohol, volumetric concentration is 85% wine by fixed blade
Essence, volumetric concentration are 90% alcohol, volumetric concentration is that 95% alcohol is dehydrated step by step, and each concentration gradient is dehydrated 45-60 minutes, then
It is put in 100% alcohol and is dehydrated twice, it is 30-45 minutes each.
Preferably, the alcoholic solution of each gradient concentration is evacuated 10- simultaneously during to leaf abscission
15 minutes.
Preferably, first blade is put under the infrared light that wavelength is 5-15 μm before dewatering and is irradiated 45-70 seconds, then put
In volumetric concentration is 80% alcohol, volumetric concentration is 85% alcohol, volumetric concentration is 90% alcohol, volumetric concentration is 95% alcohol
It is dehydrated step by step, vaned alcoholic solution will be put in every grade of dehydration and is put under the infrared light that wavelength is 5-15 μm and is irradiated
50-80 seconds, after the completion of dehydration, blade is still put under the infrared light that wavelength is 5-15 μm and is irradiated 30-60 seconds, is then placed in
It is disseminated in the sarranine solution of 0.8-1.2%.
Preferably ,+1/2 volume TO type biology of 1/2 volume absolute alcohol is successively used to the blade after the dyeing of sarranine solution
Clarifier mixed liquor and+2/3 volume TO type biology clarifier mixed liquor of 1/3 volume absolute alcohol progress transparent processing, every grade
Clearing time is 45-60 minutes；The blade after transparent processing step by step is impregnated 2 times in pure TO type biology clarifier again, often
Secondary immersion 60-90 minutes.
Preferably, in fixing step, blade is put under the infrared light that wavelength is 7-15 μm and is irradiated 45-75 seconds, then
It is put in FAA fixer and fixes, taken out after 3 hours are fixed in FAA fixer, continue to be put in the infrared light that wavelength is 7-15 μm
Lower irradiation 30-55 seconds is placed into FAA fixer later and is fixed, and after the 6th hour, taking out blade and being put in wavelength is 7-15 μm
It irradiates 30-45 seconds, finally places into fixed in FAA fixer until terminating under infrared light.
Preferably, meadowrueleaf corydalis root blade is put in after being dyed 5-15 seconds in the fast green solution that volumetric concentration is 0.8-1.2%
It takes out, washes away fast green solution with absolute alcohol, be then put in the fast green solution added with phosphoric acid and chitosan and dye again again
5-10 seconds, wherein the concentration of phosphoric acid was 0.6-0.9g/L, and the concentration of chitosan is 0.1-0.3g/L, was taken out after the completion of dyeing again
Be put in added with acetic acid volumetric concentration be 50% alcoholic solution in fixation, volumetric concentration be 50% alcohol in be added acetic acid volume
For 1/ (800-1000).
The present invention is include at least the following beneficial effects:
(1) the meadowrueleaf corydalis root blade dicing method that the present invention announces, meadowrueleaf corydalis root blade cross cut structure is clear, can be 10 to 12
Meadowrue corydalis leaf chip architecture clear in structure is cut out in it, it is permanent microscope slide sample that sample can be used with long-term preservation, can
For meadowrueleaf corydalis root medicinal material microscopical characters application, for standard market；
(2) meadowrueleaf corydalis root leaf abscission rear blade is nearly transparent, and the present invention preceding first carries out sample with sarranine solution transparent
Pre-staining, the position that embeds is more acurrate when embedding in this way, sarranine solution of the invention be with 2/3 volume absolute alcohol+
The mixed liquor of 1/3 volume TO type biology clarifier is formulated for solvent, can both carry out transparent or dyed in this way,
It is time saving and energy saving；
(3) present invention, which is added acetic acid in 50% alcohol before fast green dyeing and carries out the addition of color separation acetic acid, is conducive to point
Color, so that the fast green dyeing different parts effect of sarranine becomes apparent from；
(4) be 70% alcohol with volumetric concentration after color separation, volumetric concentration is that 85% alcohol is further processed sample, can
So that respectively organizing more three-dimensional in meadowrueleaf corydalis root blade, effect is more prominent after dyeing, it is easier to observation comparison；
(5) in fast green dyeing and then that piece is sequentially placed into absolute alcohol, 1/2TO type biology clarifier+1/2 is anhydrous
In alcohol blend, every grade stop 3-10 seconds；Finally the slide for being loaded with meadowrueleaf corydalis root blade is put into TO type biology clarifier and is stopped
It stays 5-10 minutes, this step is in triplicate, it is therefore an objective to completely remove and remain in blade and slide waxing；
(6) since the absorption spectrum of the living matters such as water, amino acid in blade is concentrated mainly on infrared region, so with red
Blade is irradiated in outside line, can promote the generation of the living matters resonance effects such as blade inner cell, increase the activity and elasticity of enzyme, in turn
So that the substance in extraneous solution is easier to enter in cell, therefore, in fixed stage and water smoking, with infrared ray to sample
Piece, which carries out processing, can promote effect that is fixed and being dehydrated, accelerate process；
(7) phosphoric acid being added after fast green dyeing and chitosan being redyed, phosphoric acid and chitosan collective effect can improve
The dye-uptake of leaf fiber, then with the further fixation of acetic acid, so that each tissue contours of blade are apparent obvious.
Further advantage, target and feature of the invention will be partially reflected by the following instructions, and part will also be by this
The research and practice of invention and be understood by the person skilled in the art.
Detailed description of the invention
Fig. 1 is the paraffin section figure of control group of the invention；
Fig. 2 is the paraffin section figure of the embodiment of the present invention 1；
Fig. 3 is the paraffin section figure of the embodiment of the present invention 6.
Specific embodiment
Present invention will be described in further detail below with reference to the accompanying drawings, to enable those skilled in the art referring to specification text
Word can be implemented accordingly.
It should be noted that experimental method described in following embodiments is unless otherwise specified conventional method, institute
Reagent and material are stated, unless otherwise specified, is commercially obtained.
Embodiment 1:
(1) fixed: fresh meadowrueleaf corydalis root blade adopt it is lower after with tap water wash down surface floating dust, blot blade surface moisture.So
Vertical vane middle arteries are crosscutting afterwards, and every is about 10mm, wide 5mm, are put into the FAA fixer that volumetric concentration is 70% alcohol
24 hours are fixed, FAA fixer is 70% alcohol by volumetric concentration: glacial acetic acid: formaldehyde volume ratio is formulated for 90:5:5.
(2) it is dehydrated: material being taken out from FAA fixer, filter paper blots extra FAA fixer, is put into rubber stopper
In flat bottom glass pipe, it is in 80% alcohol that volumetric concentration, which is added, and volumetric concentration is that the additional amount of 80% alcohol is material volume
It 20 times or more, vacuum suction 10 minutes, stops 45 minutes；Pouring out volumetric concentration is 80% alcohol, and it is 85% that volumetric concentration, which is added,
Alcohol vacuum suction 10 minutes, is placed 45 minutes；Pouring out volumetric concentration is 85% alcohol, and addition volumetric concentration is 90% alcohol,
It vacuum suction 15 minutes, places 60 minutes；Pouring out volumetric concentration is 90% alcohol, and addition volumetric concentration is 95% alcohol, vacuum
Pumping 15 minutes is placed 60 minutes；Pouring out volumetric concentration is 95% alcohol, and 100% alcohol is added, vacuum suction 15 minutes, puts
It sets 30 minutes；100% alcohol is replaced, vacuum suction 15 minutes, is placed 45 minutes.
(4) transparent: to pour out absolute alcohol, before transparent, it is 1% that dewatered meadowrueleaf corydalis root blade, which is put in volumetric concentration,
Sarranine solution disseminate 11h, wherein volumetric concentration be 1% sarranine solution be with+1/3 volume TO type of 2/3 volume absolute alcohol
The mixed liquor of biological clarifier is formulated for solvent；Sarranine solution is poured out ,+1/2 volume TO type of 1/2 volume absolute alcohol is added
The mixed liquor of biological clarifier vacuum suction 15 minutes, is placed 45 minutes；Pour out+1/2 volume TO type of 1/2 volume absolute alcohol
The mixed liquor of+2/3 volume TO type biology clarifier of 1/3 volume absolute alcohol, vacuum suction is added in the mixed liquor of biological clarifier
It 15 minutes, places 55 minutes；The mixed liquor for pouring out+2/3 volume TO type biology clarifier of 1/3 volume absolute alcohol, is added pure TO
Type biology clarifier vacuum suction 15 minutes, is placed 75 minutes；TO type biology clarifier is poured out, it is transparent to change a TO type biology
Agent vacuum suction 15 minutes, is placed 75 minutes.
(5) waxdip: the small wax stone of solid is added into the glass tube of muck coptis blade for first time waxdip, and additional amount is TO type
The 1/3 of biological clarifier volume, is stoppered bottle stopper, is put into 45 DEG C of insulating boxs, and same amount of wax is added in row again after wax thawing
Block, this step totally 5 times, until wax no longer melts, last time plus wax open bottle stopper, and TO clarifier is allowed to volatilize as far as possible,
This process needs 3 days.Second of waxdip pours into blade in crucible together with wax, then crucible is moved in 60 DEG C of insulating boxs,
The wax of solidification will be poured out after all dissolving together with blade, TO clarifier, and the paraffin that molten point is 56 DEG C has been melted in change, 60
4h is placed in DEG C insulating box；Then it changes to again and has melted the paraffin that molten point is 58 DEG C, be after 2h is placed in 60 DEG C of insulating boxs
It can embed.
(6) it embeds: taking out the crucible for filling meadowrueleaf corydalis root blade from insulating box, Label clip is gone out to be placed on embedded box on one side, bottom
After portion's wax slightly coagulates, wax is coagulated on dissecting needle slightly hot carton surface of scratching on alcolhol burner, and the material in crucible is dialled in embedding immediately
Box, and material is put well according to slice direction, stand cooling.It is put into basin after surface solidification, is allowed to sink under water 30 points
Zhong Hou, takes out embedded box, and room temperature is dried, is in store for.
(7) slice is with exhibition piece: embedded box being fixed on slicer, slice thickness is mixed up and is sliced for 6-10 μm, used
Little finger applies the bonding die agent of minute quantity gelatin on glass slide, and glass slide is placed on the warm platform of 50 ± 1 DEG C of exhibitions, 3- is added on slide
5 drop distilled water take the wax band of coverslip length 1/5-2/5 to be placed on the water surface of glass slide and are allowed to open and flat rapidly, and blotting paper sucks
Excessive moisture is placed on Zhan Wentai again, is marked, and being put into after toasting 2 days in 45 DEG C of baking ovens can just carry out in next step.
(8) dewax: piece that will be dyed is sequentially placed into 3 bottles of TO type biology clarifiers, every grade 3 minutes, slough paraffin.
(9) it dyes: piece after dewaxing is sequentially placed into+1/2 volume absolute alcohol of 1/2 volume TO type biology clarifier
Mixed liquor, absolute alcohol, absolute alcohol, volumetric concentration are 95% alcohol, volumetric concentration is 85% alcohol, volumetric concentration 70%
Rehydration in alcohol, every grade 3 seconds.It is color separation 10 seconds in 50% alcohol that piece, which is put into volumetric concentration, again, and volumetric concentration is 50% wine
Being added in smart solution has acetic acid, wherein acetic acid: the volume ratio of 50% alcohol is 1:1000.Then piece is sequentially placed into volume
Concentration is 70% alcohol, volumetric concentration is in 85% alcohol, every grade 7 seconds.Finally by piece be put into volumetric concentration be 1% it is fast green
It is dyed in solution 15 seconds, it is solvent that the fast green solution that wherein volumetric concentration is 1%, which is the alcoholic solution for being 95% with volumetric concentration,
It is formulated.After the completion of dyeing, piece is put into absolute alcohol, rinses the coloring agent on piece, this grade stops 10 seconds；Again will
Piece is put into absolute alcohol, is stopped 10 seconds；Piece is put into+1/2 volume of 1/2 volume TO type biology clarifier without watery wine later
In the mixed liquor of essence, stop 3 seconds；Finally piece is put into TO type biology clarifier, is stopped 5 minutes, this step is in triplicate.
(10) mounting: piece is lain on paper, wipes the remaining TO type biology clarifier in lower part, is dripped 2-3 above and is added dropwise
It puts on airs natural gum, coverslip dries on alcolhol burner, under carefully covering from side, avoids the occurrence of bubble, be put into 45 DEG C of baking ovens, dries
It can be taken off, post label, infuse the information such as name title material, collecting location, time on label.
Embodiment 2:
(1) fixed: fresh meadowrueleaf corydalis root blade adopt it is lower after with tap water wash down surface floating dust, blot blade surface moisture.So
Vertical vane middle arteries are crosscutting afterwards, and every is about 5mm, wide 3mm, are put into the FAA fixer that volumetric concentration is 70% alcohol
24 hours are fixed, FAA fixer is 70% alcohol by volumetric concentration: glacial acetic acid: formaldehyde volume ratio is formulated for 90:5:5.
(2) it is dehydrated: material being taken out from FAA fixer, filter paper blots extra FAA fixer, is put into rubber stopper
In flat bottom glass pipe, it is in 80% alcohol that volumetric concentration, which is added, and volumetric concentration is that the additional amount of 80% alcohol is material volume
It 20 times or more, vacuum suction 15 minutes, stops 55 minutes；Pouring out volumetric concentration is 80% alcohol, and it is 85% that volumetric concentration, which is added,
Alcohol vacuum suction 15 minutes, is placed 60 minutes；Pouring out volumetric concentration is 85% alcohol, and addition volumetric concentration is 90% alcohol,
It vacuum suction 15 minutes, places 45 minutes；Pouring out volumetric concentration is 90% alcohol, and addition volumetric concentration is 95% alcohol, vacuum
Pumping 15 minutes is placed 45 minutes；Pouring out volumetric concentration is 95% alcohol, and 100% alcohol is added, vacuum suction 15 minutes, puts
It sets 45 minutes；100% alcohol is replaced, vacuum suction 15 minutes, is placed 30 minutes.
(4) transparent: to pour out absolute alcohol, before transparent, dewatered meadowrueleaf corydalis root blade, which is put in volumetric concentration, is
1.2% sarranine solution disseminates 8h, and the sarranine solution that wherein volumetric concentration is 1.2% is with+1/3 body of 2/3 volume absolute alcohol
The mixed liquor of product TO type biology clarifier is formulated for solvent；Sarranine solution is poured out ,+1/2 body of 1/2 volume absolute alcohol is added
The mixed liquor of product TO type biology clarifier vacuum suction 10 minutes, is placed 50 minutes；Pour out+1/2 body of 1/2 volume absolute alcohol
The mixed liquor of product TO type biology clarifier, is added the mixed liquor of+2/3 volume TO type biology clarifier of 1/3 volume absolute alcohol, very
It empty pump gas 10 minutes, places 60 minutes；The mixed liquor for pouring out+2/3 volume TO type biology clarifier of 1/3 volume absolute alcohol, adds
Enter pure TO type biology clarifier, vacuum suction 10 minutes, places 60 minutes；TO type biology clarifier is poured out, it is raw to change a TO type
Object clarifier vacuum suction 10 minutes, is placed 90 minutes.
(5) waxdip: the small wax stone of solid is added into the glass tube of muck coptis blade for first time waxdip, and additional amount is TO type
The 1/3 of biological clarifier volume, is stoppered bottle stopper, is put into 45 DEG C of insulating boxs, and same amount of wax is added in row again after wax thawing
Block, this step totally 3 times, until wax no longer melts, last time plus wax open bottle stopper, and TO clarifier is allowed to volatilize as far as possible,
This process needs 2 days.Second of waxdip pours into blade in crucible together with wax, then crucible is moved in 60 DEG C of insulating boxs,
The wax of solidification will be poured out after all dissolving together with blade, TO clarifier, and the paraffin that molten point is 56 DEG C has been melted in change, 60
4h is placed in DEG C insulating box；Then it changes to again and has melted the paraffin that molten point is 58 DEG C, be after 2h is placed in 60 DEG C of insulating boxs
It can embed.
(6) it embeds: taking out the crucible for filling meadowrueleaf corydalis root blade from insulating box, Label clip is gone out to be placed on embedded box on one side, bottom
After portion's wax slightly coagulates, wax is coagulated on dissecting needle slightly hot carton surface of scratching on alcolhol burner, and the material in crucible is dialled in embedding immediately
Box, and material is put well according to slice direction, stand cooling.It is put into basin after surface solidification, is allowed to sink under water 30 points
Zhong Hou, takes out embedded box, and room temperature is dried, is in store for.
(7) slice is with exhibition piece: embedded box being fixed on slicer, slice thickness is mixed up and is sliced for 6-10 μm, used
Little finger applies the bonding die agent of minute quantity gelatin on glass slide, and glass slide is placed on the warm platform of 50 ± 1 DEG C of exhibitions, 3- is added on slide
5 drop distilled water take the wax band of coverslip length 1/5-2/5 to be placed on the water surface of glass slide and are allowed to open and flat rapidly, and blotting paper sucks
Excessive moisture is placed on Zhan Wentai again, is marked, and being put into after toasting 1 day in 45 DEG C of baking ovens can just carry out in next step.
(8) dewax: piece that will be dyed is sequentially placed into 3 bottles of TO type biology clarifiers, every grade 5 minutes, slough paraffin.
(9) it dyes: piece after dewaxing is sequentially placed into+1/2 volume absolute alcohol of 1/2 volume TO type biology clarifier
Mixed liquor, absolute alcohol, absolute alcohol, volumetric concentration are 95% alcohol, volumetric concentration is 85% alcohol, volumetric concentration 70%
Rehydration in alcohol, every grade 5 seconds.It is color separation 3 seconds in 50% alcohol that piece, which is put into volumetric concentration, again, and volumetric concentration is 50% alcohol
Being added in solution has acetic acid, wherein acetic acid: the volume ratio of 50% alcohol is 1:800.Then piece is sequentially placed into volumetric concentration
Be in 85% alcohol for 70% alcohol, volumetric concentration, every grade 10 seconds.Finally by piece be put into volumetric concentration be 1.2% it is fast green
It is dyed in solution 5 seconds, it is solvent that the fast green solution that wherein volumetric concentration is 1.2%, which is the alcoholic solution for being 95% with volumetric concentration,
It is formulated.After the completion of dyeing, piece is put into absolute alcohol, rinses the coloring agent on piece, this grade stops 8 seconds；Again will
Piece is put into absolute alcohol, is stopped 3 seconds；Piece is put into+1/2 volume of 1/2 volume TO type biology clarifier without watery wine later
In the mixed liquor of essence, stop 10 seconds；Finally piece is put into TO type biology clarifier, is stopped 10 minutes, this step repeats three
It is secondary.
(10) mounting: piece is lain on paper, wipes the remaining TO type biology clarifier in lower part, is dripped 2-3 above and is added dropwise
It puts on airs natural gum, coverslip dries on alcolhol burner, under carefully covering from side, avoids the occurrence of bubble, be put into 45 DEG C of baking ovens, dries
It can be taken off, post label, infuse the information such as name title material, collecting location, time on label.
Embodiment 3:
(1) fixed: fresh meadowrueleaf corydalis root blade adopt it is lower after with tap water wash down surface floating dust, blot blade surface moisture.So
Vertical vane middle arteries are crosscutting afterwards, and every is about 8mm, wide 4mm, are put into the FAA fixer that volumetric concentration is 70% alcohol
24 hours are fixed, FAA fixer is 70% alcohol by volumetric concentration: glacial acetic acid: formaldehyde volume ratio is formulated for 90:5:5.
(2) it is dehydrated: material being taken out from FAA fixer, filter paper blots extra FAA fixer, is put into rubber stopper
In flat bottom glass pipe, it is in 80% alcohol that volumetric concentration, which is added, and volumetric concentration is that the additional amount of 80% alcohol is material volume
It 20 times or more, vacuum suction 15 minutes, stops 60 minutes；Pouring out volumetric concentration is 80% alcohol, and it is 85% that volumetric concentration, which is added,
Alcohol vacuum suction 15 minutes, is placed 50 minutes；Pouring out volumetric concentration is 85% alcohol, and addition volumetric concentration is 90% alcohol,
It vacuum suction 15 minutes, places 55 minutes；Pouring out volumetric concentration is 90% alcohol, and addition volumetric concentration is 95% alcohol, vacuum
Pumping 15 minutes is placed 55 minutes；Pouring out volumetric concentration is 95% alcohol, and 100% alcohol is added, vacuum suction 15 minutes, puts
It sets 40 minutes；100% alcohol is replaced, vacuum suction 15 minutes, is placed 40 minutes.
(4) transparent: to pour out absolute alcohol, before transparent, dewatered meadowrueleaf corydalis root blade, which is put in volumetric concentration, is
0.8% sarranine solution disseminates 12h, and the sarranine solution that wherein volumetric concentration is 0.8% is with+1/3 body of 2/3 volume absolute alcohol
The mixed liquor of product TO type biology clarifier is formulated for solvent；Sarranine solution is poured out ,+1/2 body of 1/2 volume absolute alcohol is added
The mixed liquor of product TO type biology clarifier vacuum suction 15 minutes, is placed 60 minutes；Pour out+1/2 body of 1/2 volume absolute alcohol
The mixed liquor of product TO type biology clarifier, is added the mixed liquor of+2/3 volume TO type biology clarifier of 1/3 volume absolute alcohol, very
It empty pump gas 15 minutes, places 45 minutes；The mixed liquor for pouring out+2/3 volume TO type biology clarifier of 1/3 volume absolute alcohol, adds
Enter pure TO type biology clarifier, vacuum suction 15 minutes, places 90 minutes；TO type biology clarifier is poured out, it is raw to change a TO type
Object clarifier vacuum suction 15 minutes, is placed 60 minutes.
(5) waxdip: the small wax stone of solid is added into the glass tube of muck coptis blade for first time waxdip, and additional amount is TO type
The 1/3 of biological clarifier volume, is stoppered bottle stopper, is put into 45 DEG C of insulating boxs, and same amount of wax is added in row again after wax thawing
Block, this step totally 4 times, until wax no longer melts, last time plus wax open bottle stopper, and TO clarifier is allowed to volatilize as far as possible,
This process needs 2 days.Second of waxdip pours into blade in crucible together with wax, then crucible is moved in 60 DEG C of insulating boxs,
The wax of solidification will be poured out after all dissolving together with blade, TO clarifier, and the paraffin that molten point is 56 DEG C has been melted in change, 60
4h is placed in DEG C insulating box；Then it changes to again and has melted the paraffin that molten point is 58 DEG C, be after 2h is placed in 60 DEG C of insulating boxs
It can embed.
(6) it embeds: taking out the crucible for filling meadowrueleaf corydalis root blade from insulating box, Label clip is gone out to be placed on embedded box on one side, bottom
After portion's wax slightly coagulates, wax is coagulated on dissecting needle slightly hot carton surface of scratching on alcolhol burner, and the material in crucible is dialled in embedding immediately
Box, and material is put well according to slice direction, stand cooling.It is put into basin after surface solidification, is allowed to sink under water 30 points
Zhong Hou, takes out embedded box, and room temperature is dried, is in store for.
(7) slice is with exhibition piece: embedded box being fixed on slicer, slice thickness is mixed up and is sliced for 6-10 μm, used
Little finger applies the bonding die agent of minute quantity gelatin on glass slide, and glass slide is placed on the warm platform of 50 ± 1 DEG C of exhibitions, 3- is added on slide
5 drop distilled water take the wax band of coverslip length 1/5-2/5 to be placed on the water surface of glass slide and are allowed to open and flat rapidly, and blotting paper sucks
Excessive moisture is placed on Zhan Wentai again, is marked, and being put into after toasting 1 day in 45 DEG C of baking ovens can just carry out in next step.
(8) dewax: piece that will be dyed is sequentially placed into 3 bottles of TO type biology clarifiers, every grade 2-5 minutes, slough stone
Wax.
(9) it dyes: piece after dewaxing is sequentially placed into+1/2 volume absolute alcohol of 1/2 volume TO type biology clarifier
Mixed liquor, absolute alcohol, absolute alcohol, volumetric concentration are 95% alcohol, volumetric concentration is 85% alcohol, volumetric concentration 70%
Rehydration in alcohol, every grade 4 seconds.It is color separation 6 seconds in 50% alcohol that piece, which is put into volumetric concentration, again, and volumetric concentration is 50% alcohol
Being added in solution has acetic acid, wherein acetic acid: the volume ratio of 50% alcohol is 1:900.Then piece is sequentially placed into volumetric concentration
Be in 85% alcohol for 70% alcohol, volumetric concentration, every grade 3 seconds.Finally by piece be put into volumetric concentration be 0.8% it is fast green molten
It is dyed in liquid 10 seconds, it is solvent that the fast green solution that wherein volumetric concentration is 0.8%, which is the alcoholic solution for being 95% with volumetric concentration,
It is formulated.After the completion of dyeing, piece is put into absolute alcohol, rinses the coloring agent on piece, this grade stops 3 seconds；Again will
Piece is put into absolute alcohol, is stopped 7 seconds；Piece is put into+1/2 volume of 1/2 volume TO type biology clarifier without watery wine later
In the mixed liquor of essence, stop 7 seconds；Finally piece is put into TO type biology clarifier, is stopped 6 minutes, this step is in triplicate.
(10) mounting: piece is lain on paper, wipes the remaining TO type biology clarifier in lower part, is dripped 2-3 above and is added dropwise
It puts on airs natural gum, coverslip dries on alcolhol burner, under carefully covering from side, avoids the occurrence of bubble, be put into 45 DEG C of baking ovens, dries
It can be taken off, post label, infuse the information such as name title material, collecting location, time on label.
Embodiment 4:
On the basis of embodiment 1, in fixing step, blade is put under the infrared light that wavelength is 7-15 μm and irradiates 60
Second, then be put in FAA fixer and fix, it is taken out after 3 hours are fixed in FAA fixer, continues to be put in wavelength to be 7-15 μm
It is irradiated 45 seconds under infrared light, places into FAA fixer and fix later, after the 6th hour, taking out blade and being put in wavelength is 7-15 μm
Infrared light under irradiate 30 seconds, finally place into FAA fixer and fix 12 hours；
In dehydration, material is taken out from FAA fixer, filter paper blots extra FAA fixer, and blade is put in
It irradiates 70 seconds, is put into the flat bottom glass pipe with rubber stopper under the infrared light that wavelength is 5-15 μm, it is 80% that volumetric concentration, which is added,
It is dehydrated in alcohol, vaned alcoholic solution will be put during the dehydration process and be put under the infrared lamp that wavelength is 5-15 μm and irradiate 50
Second；Pouring out volumetric concentration is 80% alcohol, and addition volumetric concentration is 85% dehydration of alcohol, will be put during the dehydration process vaned
Alcoholic solution, which is put under the infrared lamp that wavelength is 5-15 μm, to be irradiated 80 seconds；Pouring out volumetric concentration is 85% alcohol, and volume is added
Concentration is 90% dehydration of alcohol, will put vaned alcoholic solution during the dehydration process and is put in the infrared lamp that wavelength is 5-15 μm
Lower irradiation 60 seconds；Pouring out volumetric concentration is 90% alcohol, and addition volumetric concentration is 95% dehydration of alcohol, will be put during the dehydration process
Vaned alcoholic solution, which is put under the infrared lamp that wavelength is 5-15 μm, to be irradiated 50 seconds；Pouring out volumetric concentration is 95% alcohol,
100% dehydration of alcohol is added, vaned alcoholic solution will be put during the dehydration process and be put in the infrared lamp that wavelength is 5-15 μm
Lower irradiation 50 seconds；100% dehydration of alcohol is replaced, will put vaned alcoholic solution during the dehydration process and be put in wavelength is 5-
It is irradiated 50 seconds under 15 μm of infrared lamp.
Before transparent, first blade be still put under the infrared lamp that wavelength is 5-15 μm and irradiated 30 seconds, is then placed in
It is disseminated in sarranine solution.
In staining procedure, meadowrueleaf corydalis root blade is put in after being dyed 10 seconds in fast green solution and is taken out, washed away with absolute alcohol
Then fast green solution is put in the fast green solution added with phosphoric acid and chitosan again and dyes 5 seconds again, wherein the concentration of phosphoric acid is
0.8g/L, the concentration of chitosan are 0.2g/L, and it is that 50% alcohol is molten that the volumetric concentration being put in added with acetic acid is taken out after the completion of dyeing
Fixation in liquid, volumetric concentration are that the volume of addition acetic acid in 50% alcohol is 1/1000.
Embodiment 5:
On the basis of embodiment 1, in fixing step, blade is put under the infrared lamp that wavelength is 7-15 μm and is irradiated
45 seconds, then be put in FAA fixer and fix, it is taken out after 3 hours are fixed in FAA fixer, continues to be put in wavelength to be 7-15 μm
Infrared lamp under irradiate 30 seconds, place into FAA fixer later fixed, after the 6th hour, taking out blade and being put in wavelength is 7-
It is irradiated 45 seconds under 15 μm of infrared lamp, finally places into FAA fixer and fix 12 hours；
In dehydration, material is taken out from FAA fixer, filter paper blots extra FAA fixer, and blade is put in
It irradiates 45 seconds, is put into the flat bottom glass pipe with rubber stopper under the infrared lamp that wavelength is 5-15 μm, volumetric concentration, which is added, is
It is dehydrated in 80% alcohol, vaned alcoholic solution will be put during the dehydration process and be put under the infrared lamp that wavelength is 5-15 μm and shone
It penetrates 80 seconds；Pouring out volumetric concentration is 80% alcohol, and addition volumetric concentration is 85% dehydration of alcohol, will be placed with leaf during the dehydration process
The alcoholic solution of piece, which is put under the infrared lamp that wavelength is 5-15 μm, to be irradiated 50 seconds；Pouring out volumetric concentration is 85% alcohol, is added
Volumetric concentration is 90% dehydration of alcohol, and will put vaned alcoholic solution during the dehydration process and be put in wavelength is 5-15 μm infrared
It is irradiated 50 seconds under light lamp；Pouring out volumetric concentration is 90% alcohol, and addition volumetric concentration is 95% dehydration of alcohol, during the dehydration process
Vaned alcoholic solution will be put and be put under the infrared lamp that wavelength is 5-15 μm and irradiated 80 seconds；Pouring out volumetric concentration is 95% wine
100% dehydration of alcohol is added in essence, will put vaned alcoholic solution during the dehydration process and is put in the infrared light that wavelength is 5-15 μm
It is irradiated 80 seconds under lamp；100% dehydration of alcohol is replaced, vaned alcoholic solution will be put during the dehydration process is put in wavelength and be
It is irradiated 80 seconds under 5-15 μm of infrared lamp.
Before transparent, first blade be still put under the infrared lamp that wavelength is 5-15 μm and irradiated 60 seconds, is then placed in
It is disseminated in sarranine solution.
In staining procedure, meadowrueleaf corydalis root blade is put in after being dyed 5 seconds in fast green solution and is taken out, washed away with absolute alcohol solid
Green solution, is then put in the fast green solution added with phosphoric acid and chitosan again and dyes again 10 seconds, and the volumetric concentration of phosphoric acid is
0.9g/L, the concentration of chitosan are 0.3g/L, and it is that 50% alcohol is molten that the volumetric concentration being put in added with acetic acid is taken out after the completion of dyeing
Fixation in liquid, volumetric concentration are that the volume of addition acetic acid in 50% alcohol is 1/800.
Embodiment 6:
On the basis of embodiment 1, in fixing step, blade is put under the infrared lamp that wavelength is 7-15 μm and is irradiated
75 seconds, then be put in FAA fixer and fix, it is taken out after 3 hours are fixed in FAA fixer, continues to be put in wavelength to be 7-15 μm
Infrared lamp under irradiate 55 seconds, place into FAA fixer later fixed, after the 6th hour, taking out blade and being put in wavelength is 7-
It is irradiated 40 seconds under 15 μm of infrared lamp, finally places into FAA fixer and fix 12 hours；
In dehydration, material is taken out from FAA fixer, filter paper blots extra FAA fixer, and blade is put in
It irradiates 55 seconds, is put into the flat bottom glass pipe with rubber stopper under the infrared lamp that wavelength is 5-15 μm, volumetric concentration, which is added, is
It is dehydrated in 80% alcohol, vaned alcoholic solution will be put during the dehydration process and be put under the infrared lamp that wavelength is 5-15 μm and shone
It penetrates 60 seconds；Pouring out volumetric concentration is 80% alcohol, and addition volumetric concentration is 85% dehydration of alcohol, will be placed with leaf during the dehydration process
The alcoholic solution of piece, which is put under the infrared lamp that wavelength is 5-15 μm, to be irradiated 65 seconds；Pouring out volumetric concentration is 85% alcohol, is added
Volumetric concentration is 90% dehydration of alcohol, and will put vaned alcoholic solution during the dehydration process and be put in wavelength is 5-15 μm infrared
It is irradiated 80 seconds under light lamp；Pouring out volumetric concentration is 90% alcohol, and addition volumetric concentration is 95% dehydration of alcohol, during the dehydration process
Vaned alcoholic solution will be put and be put under the infrared lamp that wavelength is 5-15 μm and irradiated 70 seconds；Pouring out volumetric concentration is 95% wine
100% dehydration of alcohol is added in essence, will put vaned alcoholic solution during the dehydration process and is put in the infrared light that wavelength is 5-15 μm
It is irradiated 55 seconds under lamp；100% dehydration of alcohol is replaced, vaned alcoholic solution will be put during the dehydration process is put in wavelength and be
It is irradiated 60 seconds under 5-15 μm of infrared lamp.
Before transparent, first blade be still put under the infrared lamp that wavelength is 5-15 μm and irradiated 45 seconds, is then placed in
It is disseminated in sarranine solution.
In staining procedure, meadowrueleaf corydalis root blade is put in the fast green solution that volumetric concentration is 0.8-1.2% and is dyed 15 seconds
After take out, wash away fast green solution with absolute alcohol, be then put in the fast green solution added with phosphoric acid and chitosan and contaminate again again
Color 8 seconds, the concentration of phosphoric acid was 0.6g/L, and the concentration of chitosan is 0.1g/L, took out the body being put in added with acetic acid after the completion of dyeing
Product concentration is fixation in 50% alcoholic solution, and volumetric concentration is that the volume of addition acetic acid in 50% alcohol is 1/900.
In order to illustrate effect of the invention, applicant use conventional paraffin section method make the paraffin section of blade as
Three, is put in Lycra biomicroscope by control group, then blade paraffin section made from Example 1 and embodiment 6 respectively
DM2000 observation is taken pictures, and respectively obtains Fig. 1, Fig. 2 and Fig. 3, as seen from the figure, the paraffin section texture of Fig. 3 is clear-cut
Clearly, it can be used for identifying the true and false of medicinal material；Fig. 2 takes second place；The root tissue of Fig. 1 is less clear, and Fig. 1 cannot be used for identifying medicine
Material.
Although the embodiments of the present invention have been disclosed as above, but its is not only in the description and the implementation listed
With it can be fully applied to various fields suitable for the present invention, for those skilled in the art, can be easily
Realize other modification, therefore without departing from the general concept defined in the claims and the equivalent scope, the present invention is simultaneously unlimited
In specific details and legend shown and described herein.
Claims (9)
1. a kind of paraffin section method of meadowrueleaf corydalis root blade, including sample is fixed, is dehydrated, is transparent, waxdip, embedding, slice and exhibition
Piece, dewaxing, dyeing and mounting, which is characterized in that
Before transparent, dewatered meadowrueleaf corydalis root blade is put in the sarranine solution that volumetric concentration is 0.8-1.2% and disseminates 8-
12h；
The following steps are included: in staining procedure, the slide that will be loaded with meadowrueleaf corydalis root blade carries out rehydration for the dyeing, after rehydration again
It is put in alcoholic solution and carries out color separation, while the vinegar that volume ratio is 1/ (800-1000) being added into the alcoholic solution for color separation
Acid is finally putting into the fast green solution that volumetric concentration is 0.8-1.2% and dyes 5-15 seconds；
In fixing step, blade is put under the infrared light that wavelength is 7-15 μm and is irradiated 45-75 seconds, then is put in FAA fixer
Middle fixation is taken out after fixing 3 hours in FAA fixer, is continued to be put under the infrared light that wavelength is 7-15 μm and is irradiated 30-55
Second, it places into FAA fixer and fixes later, after the 6th hour, take out blade and be put under the infrared light that wavelength is 7-15 μm and irradiates
It 30-45 seconds, finally places into fixed in FAA fixer until terminating.
2. the paraffin section method of meadowrueleaf corydalis root blade as described in claim 1, which is characterized in that the volumetric concentration is 0.8-
1.2% sarranine solution be with the mixed liquor of+1/3 volume TO type biology clarifier of 2/3 volume absolute alcohol be solvent prepare and
At, the fast green solution that the volumetric concentration is 0.8-1.2% be the alcoholic solution for being 95% with volumetric concentration be solvent prepare and
At.
3. the paraffin section method of meadowrueleaf corydalis root blade as described in claim 1, which is characterized in that before dyeing, will dewax
The slide for being loaded with meadowrueleaf corydalis root blade afterwards is sequentially placed into+1/2 volume absolute alcohol mixed liquor of 1/2 volume type biology clarifier, nothing
Water-alcohol, absolute alcohol, volumetric concentration are 95% alcohol, volumetric concentration is 85% alcohol, volumetric concentration is multiple in 70% alcohol
Water, every grade 3-5 seconds；It places into 50% alcohol color separation 3-10 seconds；Being sequentially placed into volumetric concentration again later is 70% alcohol, volume
Concentration be 85% alcohol in be further processed, every grade 3-10 seconds.
4. the paraffin section method of meadowrueleaf corydalis root blade as described in claim 1, which is characterized in that will be carried after fast green solution dyeing
There is the slide of meadowrueleaf corydalis root blade to be put in be put into absolute alcohol and wash away coloring agent, this grade stops 3-10 seconds；Then it is sequentially placed into nothing
In water-alcohol ,+1/2 absolute alcohol mixed liquor of 1/2TO type biology clarifier, every grade stop 3-10 seconds；Meadowrueleaf corydalis root will be finally loaded with
The slide of blade is put into TO type biology clarifier and stops 5-10 minutes, this step is in triplicate.
5. the paraffin section method of meadowrueleaf corydalis root blade as described in claim 1, which is characterized in that will by fixed blade according to
Secondary process volumetric concentration is 80% alcohol, volumetric concentration is 85% alcohol, volumetric concentration is 90% alcohol, volumetric concentration 95%
Alcohol is dehydrated step by step, and each concentration gradient is dehydrated 45-60 minutes, then is put in 100% alcohol and is dehydrated twice, and each 30-45 points
Clock.
6. the paraffin section method of meadowrueleaf corydalis root blade as claimed in claim 5, which is characterized in that the alcohol of each gradient concentration
Solution is evacuated 10-15 minutes simultaneously during to leaf abscission.
7. the paraffin section method of meadowrueleaf corydalis root blade as claimed in claim 5, which is characterized in that before dewatering first by blade
It is put under the infrared light that wavelength is 5-15 μm and irradiates 45-70 seconds, then being put in volumetric concentration is 80% alcohol, volumetric concentration 85%
Alcohol, volumetric concentration are 90% alcohol, volumetric concentration is that 95% alcohol is dehydrated step by step, and leaf will be placed in every grade of dehydration
The alcoholic solution of piece, which is put under the infrared light that wavelength is 5-15 μm, to be irradiated 50-80 seconds, and after the completion of dehydration, blade is still put in wave
It is irradiated 30-60 seconds under a length of 5-15 μm of infrared light, is then placed in the sarranine solution of 0.8-1.2% and disseminates.
8. the paraffin section method of meadowrueleaf corydalis root blade as described in claim 1, which is characterized in that after the dyeing of sarranine solution
Blade successively uses+1/2 volume TO type biology clarifier mixed liquor of 1/2 volume absolute alcohol and 1/3 volume absolute alcohol+2/3
Volume TO type biology clarifier mixed liquor carries out transparent processing, and every grade of clearing time is 45-60 minutes；It again will transparent place step by step
Blade after reason impregnates 2 times in pure TO type biology clarifier, impregnates 60-90 minutes every time.
9. the paraffin section method of meadowrueleaf corydalis root blade as described in claim 1, which is characterized in that meadowrueleaf corydalis root blade is put in body
It is taken out after being dyed 5-15 seconds in the fast green solution that product concentration is 0.8-1.2%, washes away fast green solution with absolute alcohol, then put again
It is dyed again in the fast green solution added with phosphoric acid and chitosan 5-10 seconds, wherein the concentration of phosphoric acid is 0.6-0.9g/L, shell
The concentration of glycan be 0.1-0.3g/L, again dyeing after the completion of take out be put in added with acetic acid volumetric concentration be 50% alcoholic solution
Middle fixation, volumetric concentration are that the volume of addition acetic acid in 50% alcohol is 1/ (800-1000).
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| CN107702959B (en) * | 2017-10-27 | 2020-12-01 | 陕西省西安植物园 | Paraffin section method for gradient dehydration of plant material |
| CN109374376A (en) * | 2018-11-09 | 2019-02-22 | 上海市农业科学院 | A kind of slice preparation method suitable for mushroom lamella Basidium morphologic observation |
| CN109632431A (en) * | 2019-02-12 | 2019-04-16 | 北京林业大学 | A kind of simple and quick clearing method |
| 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 |
| CN110553890A (en) * | 2019-10-16 | 2019-12-10 | 甘肃农业大学 | safe and efficient paraffin slicing method for potato roots and leaves |
| CN113624580A (en) * | 2021-07-20 | 2021-11-09 | 四川农业大学 | Dyeing method of bamboo plant veins |
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| CN103630421A (en) * | 2013-12-03 | 2014-03-12 | 沈阳农业大学 | Production method for paraffin section of paeonia lactiflora mature embryo |
| CN105806682A (en) * | 2016-03-08 | 2016-07-27 | 中国环境科学研究院 | Method for preparing leguminous plant leaf paraffin section |
| CN105973673A (en) * | 2016-06-30 | 2016-09-28 | 中国林业科学研究院热带林业研究所 | Paraffin sectioning method for eucalyptus tissue |
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Application publication date: 20170315 Assignee: Guangxi Guinong Precision Farming Machinery Service Co.,Ltd. Assignor: GUANGXI BOTANICAL GARDEN OF MEDICINAL PLANTS Contract record no.: X2023980045319 Denomination of invention: Paraffin sectioning method for leaves of Yanhuanglian Granted publication date: 20190226 License type: Common License Record date: 20231101 |