CN112816296A - Hematoxylin-alkaline re-redden-picric acid staining kit and staining method thereof - Google Patents
Hematoxylin-alkaline re-redden-picric acid staining kit and staining method thereof Download PDFInfo
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- CN112816296A CN112816296A CN202011643069.8A CN202011643069A CN112816296A CN 112816296 A CN112816296 A CN 112816296A CN 202011643069 A CN202011643069 A CN 202011643069A CN 112816296 A CN112816296 A CN 112816296A
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- 238000010186 staining Methods 0.000 title claims abstract description 27
- 238000007447 staining method Methods 0.000 title abstract description 7
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims abstract description 92
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 claims abstract description 72
- OXNIZHLAWKMVMX-UHFFFAOYSA-N picric acid Chemical compound OC1=C([N+]([O-])=O)C=C([N+]([O-])=O)C=C1[N+]([O-])=O OXNIZHLAWKMVMX-UHFFFAOYSA-N 0.000 claims abstract description 49
- 239000007788 liquid Substances 0.000 claims abstract description 26
- 238000002360 preparation method Methods 0.000 claims abstract description 25
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims abstract description 21
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 20
- 239000012153 distilled water Substances 0.000 claims abstract description 17
- UKWHYYKOEPRTIC-UHFFFAOYSA-N mercury(ii) oxide Chemical compound [Hg]=O UKWHYYKOEPRTIC-UHFFFAOYSA-N 0.000 claims abstract description 14
- 238000000034 method Methods 0.000 claims abstract description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 7
- 229940103272 aluminum potassium sulfate Drugs 0.000 claims abstract description 7
- 229940101209 mercuric oxide Drugs 0.000 claims abstract description 7
- GRLPQNLYRHEGIJ-UHFFFAOYSA-J potassium aluminium sulfate Chemical compound [Al+3].[K+].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O GRLPQNLYRHEGIJ-UHFFFAOYSA-J 0.000 claims abstract description 7
- JUQPZRLQQYSMEQ-UHFFFAOYSA-N CI Basic red 9 Chemical compound [Cl-].C1=CC(N)=CC=C1C(C=1C=CC(N)=CC=1)=C1C=CC(=[NH2+])C=C1 JUQPZRLQQYSMEQ-UHFFFAOYSA-N 0.000 claims abstract 5
- 229940052223 basic fuchsin Drugs 0.000 claims abstract 5
- 239000012188 paraffin wax Substances 0.000 claims description 27
- 239000001054 red pigment Substances 0.000 claims description 12
- 238000005406 washing Methods 0.000 claims description 11
- 238000004043 dyeing Methods 0.000 claims description 8
- CTQNGGLPUBDAKN-UHFFFAOYSA-N O-Xylene Chemical compound CC1=CC=CC=C1C CTQNGGLPUBDAKN-UHFFFAOYSA-N 0.000 claims description 3
- 230000018044 dehydration Effects 0.000 claims description 3
- 238000006297 dehydration reaction Methods 0.000 claims description 3
- 230000007935 neutral effect Effects 0.000 claims description 3
- 239000001044 red dye Substances 0.000 claims description 3
- 239000011347 resin Substances 0.000 claims description 3
- 229920005989 resin Polymers 0.000 claims description 3
- 238000007789 sealing Methods 0.000 claims description 3
- 239000008399 tap water Substances 0.000 claims description 3
- 235000020679 tap water Nutrition 0.000 claims description 3
- 239000008096 xylene Substances 0.000 claims description 3
- 238000011010 flushing procedure Methods 0.000 claims description 2
- 208000031225 myocardial ischemia Diseases 0.000 description 5
- 230000000694 effects Effects 0.000 description 3
- 210000004165 myocardium Anatomy 0.000 description 3
- 230000009286 beneficial effect Effects 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- MCSXGCZMEPXKIW-UHFFFAOYSA-N 3-hydroxy-4-[(4-methyl-2-nitrophenyl)diazenyl]-N-(3-nitrophenyl)naphthalene-2-carboxamide Chemical compound Cc1ccc(N=Nc2c(O)c(cc3ccccc23)C(=O)Nc2cccc(c2)[N+]([O-])=O)c(c1)[N+]([O-])=O MCSXGCZMEPXKIW-UHFFFAOYSA-N 0.000 description 1
- 102000004506 Blood Proteins Human genes 0.000 description 1
- 108010017384 Blood Proteins Proteins 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000008081 blood perfusion Effects 0.000 description 1
- 210000003855 cell nucleus Anatomy 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 210000004177 elastic tissue Anatomy 0.000 description 1
- 230000037149 energy metabolism Effects 0.000 description 1
- 210000003743 erythrocyte Anatomy 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 230000000302 ischemic effect Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
- G01N1/31—Apparatus therefor
-
- 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)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Physics & Mathematics (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)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a hematoxylin-alkaline re-redden-picric acid staining kit and a staining method thereof. The kit comprises: harris hematoxylin solution, alkaline re-reddening solution and picric acid acetone solution; the preparation method of the Harris hematoxylin liquid comprises the following steps: adding Harris hematoxylin into absolute ethyl alcohol to obtain a solution A, adding aluminum potassium sulfate into distilled water to obtain a solution B, mixing the solution A and the solution B in a volume ratio of 1:20 to obtain a solution C, adding mercuric oxide into the solution C, standing for 12-24 hours to obtain a solution D, and adding citric acid into the solution D to obtain a Harris hematoxylin solution, wherein the Harris hematoxylin solution comprises 0.01-0.03% of Harris hematoxylin in percentage by mass; the preparation method of the alkaline reddening solution comprises the following steps: adding the basic fuchsin into distilled water to obtain a basic fuchsin liquid, wherein the basic fuchsin liquid contains 0.08-0.12% of the basic fuchsin by mass percent; the preparation method of the picric acid acetone solution comprises the following steps: the picric acid is added into acetone to prepare the picric acid-acetone liquid, and the picric acid-acetone liquid contains 0.08-0.12% of picric acid in percentage by mass. The scheme has the advantages of simple preparation method, low cost and simple use method.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a hematoxylin-alkaline re-reddening-picric acid staining kit and a staining method thereof.
Background
Myocardial ischemia (myocardial ischemia) refers to a pathological condition in which the blood perfusion of the heart is reduced, the oxygen supply of the heart is reduced, the energy metabolism of the heart muscle is abnormal, and the heart cannot work normally. With the gradual younger patients with myocardial ischemia, the early change of myocardial ischemia is pathologically diagnosed, and the observation and research of experimental myocardial ischemia materials are of great significance.
Disclosure of Invention
In order to overcome the defects in the prior art, the embodiment of the invention provides a hematoxylin-basic fuchsin-picric acid staining kit and a staining method thereof, and provides the kit with simple preparation method, low cost and simple use method.
The embodiment of the application discloses: a hematoxylin-alkaline re-reddish-picric acid staining kit comprises: harris hematoxylin solution, alkaline re-reddening solution and picric acid acetone solution; wherein:
the preparation method of the Harris hematoxylin liquid comprises the following steps: adding Harris hematoxylin into absolute ethyl alcohol to obtain a solution A, adding aluminum potassium sulfate into distilled water to be fully dissolved to obtain a solution B, mixing the solution A and the solution B in a volume ratio of 1:20 to obtain a solution C, adding mercuric oxide into the solution C, mixing, standing for 12-24 hours to obtain a solution D, adding citric acid into the solution D, and uniformly mixing to obtain the Harris hematoxylin solution, wherein the Harris hematoxylin solution comprises 0.01-0.03% by mass;
the preparation method of the alkaline reddening solution comprises the following steps: adding alkaline red pigment into distilled water to obtain the alkaline red pigment liquid, wherein the alkaline red pigment liquid contains 0.08-0.12% of the alkaline red pigment by mass percent;
the preparation method of the picric acid acetone solution comprises the following steps: adding picric acid into acetone to obtain the picric acid acetone solution, wherein the picric acid acetone solution contains 0.08-0.12% of picric acid in percentage by mass.
Specifically, the preparation method of the Harris hematoxylin solution comprises the following steps: adding 2.5 g of Harris hematoxylin into 25 ml of absolute ethyl alcohol to obtain a solution A, adding 22 ml of aluminum potassium sulfate into 500 ml of distilled water to be fully dissolved to obtain a solution B, mixing the solution A and the solution B to obtain a solution C, adding 1.25 g of mercuric oxide into the solution C, mixing, standing for 12-24 hours to obtain a solution D, adding 2 g of citric acid into the solution D, and uniformly mixing to obtain the Harris hematoxylin solution.
Specifically, the preparation method of the alkaline reddening solution comprises the following steps: 0.1 g of basic reddish was added to 100 ml of distilled water to obtain the basic reddish liquid.
Specifically, the preparation method of the picric acid acetone solution comprises the following steps: 0.1 g picric acid was added to 100 ml acetone to obtain the picric acid acetone solution.
Specifically, the alkaline reddish liquid contains 0.1% of alkaline reddish, and the picric acid acetone liquid contains 0.1% of picric acid.
Specifically, the hematoxylin-alkaline re-red-picric acid staining kit comprises: 500 ml of Harris hematoxylin solution, 100 ml of alkaline reddening solution and 100 ml of picric acid acetone solution.
This implementation still discloses: the method for staining by using the hematoxylin-alkaline re-reddish-picric acid staining kit comprises the following steps of:
step 1: paraffin section dewaxing to water washing;
step 2: immersing the paraffin section obtained in the step 1 into the Harris hematoxylin liquid for dyeing for 30-35 seconds, and then flushing with tap water for 300-305 seconds;
and step 3: immersing the paraffin section obtained in the step 2 into the alkaline reddening solution for dyeing for 180-185 seconds, and then washing with distilled water for 5-10 seconds;
and 4, step 4: washing the paraffin sections obtained in the step 3 with pure acetone for 5-10 seconds;
and 5: differentiating the paraffin sections obtained in the step 4 by adopting picric acid acetone liquid for 5-15 seconds so that no red dye liquid flows down on the paraffin sections;
step 6: carrying out pure acetone dehydration on the paraffin section obtained in the step 5 for 5-10 seconds;
and 7: and (4) sequentially carrying out xylene transparency and neutral resin sealing on the paraffin sections obtained in the step (6).
Specifically, the alkaline red recovery liquid comprises 0.1% of alkaline red recovery liquid in percentage by mass.
Specifically, the picric acid acetone solution comprises 0.1% of picric acid in percentage by mass.
Specifically, the thickness of the paraffin section is between 3 and 10 micrometers.
The invention has at least the following beneficial effects:
the hematoxylin-alkaline re-reddish-picric acid staining kit provided by the embodiment has the advantages of simple preparation method, low cost, simple staining method and obvious staining effect.
In order to make the aforementioned and other objects, features and advantages of the invention more comprehensible, preferred embodiments accompanied with figures are described in detail below.
Detailed Description
The technical solutions in the embodiments of the present invention will be described clearly and completely with reference to the following embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The hematoxylin-basic re-reddish-picric acid staining kit of the embodiment comprises: harris hematoxylin solution, alkaline re-reddening solution and picric acid acetone solution. Preferably, the liquid amount of each component is: 500 ml of Harris hematoxylin solution, 100 ml of alkaline reddening solution and 100 ml of picric acid acetone solution.
The preparation method of each component comprises the following steps:
the preparation method of the Harris hematoxylin liquid comprises the following steps: adding Harris hematoxylin into absolute ethyl alcohol to obtain a solution A, adding aluminum potassium sulfate into distilled water to be fully dissolved to obtain a solution B, mixing the solution A and the solution B in a volume ratio of 1:20 to obtain a solution C, adding mercuric oxide into the solution C, mixing, standing for 12-24 hours to obtain a solution D, adding citric acid into the solution D, and uniformly mixing to obtain the Harris hematoxylin solution, wherein the Harris hematoxylin solution comprises 0.01-0.03% by mass, preferably 0.02% by mass.
The preparation method of the alkaline reddening solution comprises the following steps: adding the basic red pigment into distilled water to obtain the basic red pigment solution, wherein the basic red pigment solution comprises 0.08-0.12% of the basic red pigment in percentage by mass.
The preparation method of the picric acid acetone solution comprises the following steps: adding picric acid into acetone to obtain the picric acid acetone solution, wherein the picric acid acetone solution contains 0.08-0.12% of picric acid in percentage by mass.
In a specific embodiment, the preparation method of the Harris hematoxylin solution comprises the following steps: adding 2.5 g of Harris hematoxylin into 25 ml of absolute ethyl alcohol to obtain a solution A, adding 22 ml of aluminum potassium sulfate into 500 ml of distilled water to be fully dissolved to obtain a solution B, mixing the solution A and the solution B to obtain a solution C, adding 1.25 g of mercuric oxide into the solution C, mixing, standing for 12-24 hours to obtain a solution D, adding 2 g of citric acid into the solution D, and uniformly mixing to obtain the Harris hematoxylin solution.
In a specific embodiment, the preparation method of the alkaline reddening solution comprises the following steps: 0.1 g of basic reddish was added to 100 ml of distilled water to obtain the basic reddish liquid.
In a specific embodiment, the preparation method of the picric acid acetone solution comprises the following steps: 0.1 g picric acid was added to 100 ml acetone to obtain the picric acid acetone solution.
Specifically, the alkaline reddening solution contains 0.1% of alkaline reddening and the picric acid acetone solution contains 0.1% of picric acid in percentage by mass, so that the dyeing effect is favorably improved.
The method for staining by adopting the hematoxylin-alkaline re-reddening-picric acid staining kit comprises the following steps:
step 1: paraffin section dewaxing to water washing;
step 2: immersing the paraffin section obtained in the step 1 into the Harris hematoxylin liquid for staining for 30-35 seconds, and then washing for 300-305 seconds by using tap water, wherein in the step, the staining is preferably performed for 30 seconds, and the washing is preferably performed for 300 seconds;
and step 3: immersing the paraffin section obtained in the step 2 into the alkaline reddening solution for dyeing for 180-185 seconds, and then washing with distilled water for 5-10 seconds, wherein the dyeing time is preferably 180 seconds in the step;
and 4, step 4: washing the paraffin sections obtained in the step 3 with pure acetone for 5-10 seconds;
and 5: differentiating the paraffin sections obtained in the step 4 by adopting the picric acid acetone solution for 5-15 seconds so that no red dye solution flows down on the paraffin sections;
step 6: carrying out pure acetone dehydration on the paraffin section obtained in the step 5 for 5-10 seconds;
and 7: and (4) sequentially carrying out xylene transparency and neutral resin sealing on the paraffin sections obtained in the step (6).
Specifically, in the step 3, the basic reddening liquid comprises 0.1% of basic reddening in percentage by mass; in the step 5, the picric acid acetone solution comprises 0.1% of picric acid by mass percentage, which is beneficial to improving the dyeing effect.
Specifically, the thickness of the paraffin section is between 3 and 10 μm, and can be properly adjusted according to actual conditions.
After the hematoxylin-alkaline re-reddening-picric acid staining kit and the staining method are adopted for staining, the anoxic and ischemic cardiac muscle, the red blood cells, the plasma protein, the elastic fiber and the collagen fiber are all bright red or deep red, the normal cardiac muscle is yellow or yellowish brown, and the cell nucleus is black blue.
The principle and the implementation mode of the invention are explained by applying specific embodiments in the invention, and the description of the embodiments is only used for helping to understand the method and the core idea of the invention; meanwhile, for a person skilled in the art, according to the idea of the present invention, there may be variations in the specific embodiments and the application scope, and in summary, the content of the present specification should not be construed as a limitation to the present invention.
Claims (10)
1. A hematoxylin-alkaline re-reddish-picric acid staining kit is characterized by comprising: harris hematoxylin solution, alkaline re-reddening solution and picric acid acetone solution; wherein:
the preparation method of the Harris hematoxylin liquid comprises the following steps: adding Harris hematoxylin into absolute ethyl alcohol to obtain a solution A, adding aluminum potassium sulfate into distilled water to be fully dissolved to obtain a solution B, mixing the solution A and the solution B in a volume ratio of 1:20 to obtain a solution C, adding mercuric oxide into the solution C, mixing, standing for 12-24 hours to obtain a solution D, adding citric acid into the solution D, and uniformly mixing to obtain the Harris hematoxylin solution, wherein the Harris hematoxylin solution comprises 0.01-0.03% by mass;
the preparation method of the alkaline reddening solution comprises the following steps: adding alkaline red pigment into distilled water to obtain the alkaline red pigment liquid, wherein the alkaline red pigment liquid contains 0.08-0.12% of the alkaline red pigment by mass percent;
the preparation method of the picric acid acetone solution comprises the following steps: adding picric acid into acetone to obtain the picric acid acetone solution, wherein the picric acid acetone solution contains 0.08-0.12% of picric acid in percentage by mass.
2. The hematoxylin-basic re-reddish-picric acid staining kit according to claim 1, wherein the preparation method of the Harris hematoxylin solution comprises: adding 2.5 g of Harris hematoxylin into 25 ml of absolute ethyl alcohol to obtain a solution A, adding 22 ml of aluminum potassium sulfate into 500 ml of distilled water to be fully dissolved to obtain a solution B, mixing the solution A and the solution B to obtain a solution C, adding 1.25 g of mercuric oxide into the solution C, mixing, standing for 12-24 hours to obtain a solution D, adding 2 g of citric acid into the solution D, and uniformly mixing to obtain the Harris hematoxylin solution.
3. The hematoxylin-alkaline re-reddish-picric acid staining kit according to claim 1, wherein the preparation method of the alkaline re-reddish solution comprises the following steps: 0.1 g of basic reddish was added to 100 ml of distilled water to obtain the basic reddish liquid.
4. The hematoxylin-basic re-reddish-picric acid staining kit according to claim 1, wherein the preparation method of the picric acid acetone solution comprises: 0.1 g picric acid was added to 100 ml acetone to obtain the picric acid acetone solution.
5. The hematoxylin-basic re-redness-picric acid staining kit according to claim 1, wherein the basic re-redness solution comprises 0.1% of basic re-redness, and the picric acid acetone solution comprises 0.1% of picric acid by mass.
6. The hematoxylin-alkaline re-reddish-picric acid staining kit of claim 1, wherein the hematoxylin-alkaline re-reddish-picric acid staining kit comprises: 500 ml of Harris hematoxylin solution, 100 ml of alkaline reddening solution and 100 ml of picric acid acetone solution.
7. A method of staining using the hematoxylin-basic re-reddish-picric acid staining kit of any one of claims 1 to 6, comprising the steps of:
step 1: paraffin section dewaxing to water washing;
step 2: immersing the paraffin section obtained in the step 1 into the Harris hematoxylin liquid for dyeing for 30-35 seconds, and then flushing with tap water for 300-305 seconds;
and step 3: immersing the paraffin section obtained in the step 2 into the alkaline reddening solution for dyeing for 180-185 seconds, and then washing with distilled water for 5-10 seconds;
and 4, step 4: washing the paraffin sections obtained in the step 3 with pure acetone for 5-10 seconds;
and 5: differentiating the paraffin sections obtained in the step 4 by adopting the picric acid acetone solution for 5-15 seconds so that no red dye solution flows down on the paraffin sections;
step 6: carrying out pure acetone dehydration on the paraffin section obtained in the step 5 for 5-10 seconds;
and 7: and (4) sequentially carrying out xylene transparency and neutral resin sealing on the paraffin sections obtained in the step (6).
8. The method according to claim 7, wherein in step 3, the basic fuchsin is 0.1% by mass.
9. The method as claimed in claim 7, wherein in step 5, picric acid 0.1% is included in the picric acid acetone solution.
10. The method according to claim 7, wherein the paraffin section has a thickness of 3-10 μm.
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| CN202011643069.8A CN112816296A (en) | 2020-12-30 | 2020-12-30 | Hematoxylin-alkaline re-redden-picric acid staining kit and staining method thereof |
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| CN202011643069.8A CN112816296A (en) | 2020-12-30 | 2020-12-30 | Hematoxylin-alkaline re-redden-picric acid staining kit and staining method thereof |
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106668026A (en) * | 2017-02-22 | 2017-05-17 | 中国人民武装警察部队后勤学院 | Application of rosolic acid to preparation of anti-hypoxic medicine |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106668026A (en) * | 2017-02-22 | 2017-05-17 | 中国人民武装警察部队后勤学院 | Application of rosolic acid to preparation of anti-hypoxic medicine |
Non-Patent Citations (3)
| Title |
|---|
| 全国卫生专业技术资格考试专家委员会: "《卫生专业技术资格考试指导 病理学与病理学技术（上册）》", 28 February 2004 * |
| 成均 等: "《现代临床内科学》", 31 March 2009 * |
| 王莉芬: "《病理学 高级医师进阶 第2版》", 31 January 2020 * |
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Application publication date: 20210518 |