CN112504777A - Method for fixing egg yolk membrane protein and application thereof - Google Patents
Method for fixing egg yolk membrane protein and application thereof Download PDFInfo
- Publication number
- CN112504777A CN112504777A CN202011155531.XA CN202011155531A CN112504777A CN 112504777 A CN112504777 A CN 112504777A CN 202011155531 A CN202011155531 A CN 202011155531A CN 112504777 A CN112504777 A CN 112504777A
- Authority
- CN
- China
- Prior art keywords
- membrane
- egg
- yolk
- yolk membrane
- egg yolk
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 108010000912 Egg Proteins Proteins 0.000 title claims abstract description 32
- 102000002322 Egg Proteins Human genes 0.000 title claims abstract description 32
- 210000002969 egg yolk Anatomy 0.000 title claims abstract description 29
- 238000000034 method Methods 0.000 title claims abstract description 25
- 108010052285 Membrane Proteins Proteins 0.000 title claims abstract description 24
- 235000013345 egg yolk Nutrition 0.000 title claims abstract description 24
- 102000018697 Membrane Proteins Human genes 0.000 title claims abstract description 20
- 210000001534 vitelline membrane Anatomy 0.000 claims abstract description 24
- 239000012528 membrane Substances 0.000 claims abstract description 16
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 15
- WMFOQBRAJBCJND-UHFFFAOYSA-M Lithium hydroxide Chemical compound [Li+].[OH-] WMFOQBRAJBCJND-UHFFFAOYSA-M 0.000 claims abstract description 14
- 235000013601 eggs Nutrition 0.000 claims abstract description 14
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 4
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 4
- 238000005507 spraying Methods 0.000 claims abstract description 3
- KMUONIBRACKNSN-UHFFFAOYSA-N potassium dichromate Chemical compound [K+].[K+].[O-][Cr](=O)(=O)O[Cr]([O-])(=O)=O KMUONIBRACKNSN-UHFFFAOYSA-N 0.000 claims description 16
- 239000012153 distilled water Substances 0.000 claims description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 229960002523 mercuric chloride Drugs 0.000 claims description 8
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 claims description 8
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 claims description 5
- 210000000969 egg white Anatomy 0.000 claims description 5
- 235000014103 egg white Nutrition 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- QLOKJRIVRGCVIM-UHFFFAOYSA-N 1-[(4-methylsulfanylphenyl)methyl]piperazine Chemical compound C1=CC(SC)=CC=C1CN1CCNCC1 QLOKJRIVRGCVIM-UHFFFAOYSA-N 0.000 claims description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 3
- 210000003278 egg shell Anatomy 0.000 claims description 3
- 239000000203 mixture Substances 0.000 claims description 3
- 239000002244 precipitate Substances 0.000 claims description 2
- 230000001954 sterilising effect Effects 0.000 claims description 2
- 239000010902 straw Substances 0.000 claims description 2
- 239000006228 supernatant Substances 0.000 claims description 2
- 108010022099 vitelline membrane proteins Proteins 0.000 claims 5
- 230000003100 immobilizing effect Effects 0.000 claims 3
- 229910052751 metal Inorganic materials 0.000 claims 1
- 239000002184 metal Substances 0.000 claims 1
- 238000004043 dyeing Methods 0.000 abstract description 6
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 abstract description 3
- 239000010931 gold Substances 0.000 abstract description 3
- 229910052737 gold Inorganic materials 0.000 abstract description 3
- 230000009286 beneficial effect Effects 0.000 abstract description 2
- QSHDDOUJBYECFT-UHFFFAOYSA-N mercury Chemical compound [Hg] QSHDDOUJBYECFT-UHFFFAOYSA-N 0.000 abstract description 2
- 229910021645 metal ion Inorganic materials 0.000 abstract description 2
- 239000002245 particle Substances 0.000 description 3
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012876 topography Methods 0.000 description 2
- 230000001133 acceleration Effects 0.000 description 1
- 238000007792 addition Methods 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000000249 desinfective effect Effects 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
Images
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
-
- 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
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Physics & Mathematics (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention discloses a method for fixing egg yolk membrane protein and application thereof. The invention relates to a method for fixing egg yolk membrane protein, which comprises the steps of processing eggs to obtain a yolk membrane, dyeing the yolk membrane by using a Golgi-Cox reagent, and blackening the yolk membrane by using LiOH. A method for observing the shapes of egg yolk membrane and membrane protein thereof comprises the steps of fixing the egg yolk membrane protein by the method, then spreading the fixed egg yolk membrane on carbon conductive gel, naturally airing, storing in dark place, spraying gold, and performing scanning analysis by an electron microscope, so that the shapes of the egg yolk membrane and the membrane protein thereof can be observed. The Golgi-Cox dyeing method is used for fixing egg yolk membranes and membrane proteins thereof, mercury metal ions in the Golgi-Cox dyeing method are beneficial to fixing membrane surface proteins, the method is simple, and experimental results of the method for observing the shapes of the egg yolk membranes and the membrane proteins thereof are quick and available.
Description
Technical Field
The invention relates to the technical field of biomedicine, in particular to a method for fixing egg yolk membrane protein and application thereof.
Background
The egg yolk membrane of the egg is mainly composed of protein, which accounts for 81.6% (w/w) of the dry weight of the yolk membrane, and a small amount of lipid and carbohydrate accounts for 7.8%, and a very small amount of impurities accounts for 0.61%. The appearance observation of the egg yolk membrane protein requires the fixation of the sample. Common chemical fixation easily changes the structure of the yolk membrane, while the sample preparation conditions in the freezing technology are harsh, and the sample preparation process is multiple and expensive.
Disclosure of Invention
The invention aims to provide a simple method for fixing egg yolk membrane protein and application thereof, aiming at the defects in the prior art.
The invention relates to a method for fixing egg yolk membrane protein, which comprises the steps of processing eggs to obtain a yolk membrane, dyeing the yolk membrane by using a Golgi-Cox reagent, and blackening the yolk membrane by using LiOH.
Further, the specific operation of processing the eggs to obtain the vitelline membrane is as follows: sterilizing whole fresh egg with 75% alcohol, breaking egg shell, pouring egg white and yolk into a beaker, sucking out egg white with a disposable straw, sucking out yolk membrane, sucking out yolk in the yolk membrane, and rinsing the yolk membrane with distilled water.
Further, the rinsed yolk membrane was stained in Golgi-Cox reagent for 2 days and 1% LiOH was blackened for one day.
Further, the Golgi-Cox reagent comprises the following components in parts by mass: 1 part of mercuric chloride, 1 part of potassium dichromate, 0.8 part of potassium chromate and 80 parts of distilled water.
Further, the Golgi-Cox reagent formulation process is as follows: firstly, 1 part of mercuric chloride and 1 part of potassium dichromate are dissolved in 64 parts of distilled water, 0.8 part of potassium chromate is dissolved in 16 parts of distilled water in a brown wide-mouth reagent bottle, the reagent bottle is shaken for more than 12 hours at room temperature, then the mercuric chloride and the potassium dichromate are uniformly mixed, the mercuric chloride and the potassium dichromate are kept overnight at the temperature of more than 4 hours or 4 ℃ by a greenhouse shaking table, and a supernatant is taken after a precipitate is deposited at the bottom of the reagent bottle, and the mixture is used for preparation or kept away from light.
A method for observing the shapes of egg yolk membrane and membrane protein thereof comprises the steps of fixing the egg yolk membrane protein by the method, then spreading the fixed egg yolk membrane on carbon conductive gel, naturally airing, storing in dark place, spraying gold, and performing scanning analysis by an electron microscope, so that the shapes of the egg yolk membrane and the membrane protein thereof can be observed.
The Golgi-Cox dyeing method is used for fixing the egg yolk membrane and the membrane protein thereof, and mercury metal ions in the Golgi-Cox dyeing method are beneficial to fixing the membrane surface protein.
Drawings
FIG. 1 is a 20000 times magnified topography of micron-sized membrane protein particles on a Golgi-Cox stained LiOH blackened egg yolk membrane sample prepared in example 1 of the present invention.
FIG. 2 is a 40000 times magnified topography of micron-sized membrane protein particles on a Golgi-Cox stained LiOH blackened egg yolk membrane sample made in example 1 of the present invention.
Detailed Description
The following are specific embodiments of the present invention and are further described with reference to the drawings, but the present invention is not limited to these embodiments.
Example 1
Disinfecting the shell of the egg with alcohol, breaking the eggshell, sucking out the egg white with a 5ml disposable rubber dropper, sucking the yolk membrane, removing the yolk, and rinsing with distilled water. Golgi-Cox staining was then performed for 2 days and blackening for 1 day. Then, the dyed yolk membrane is taken out, laid on carbon conductive gel for natural drying, and sprayed with gold for scanning analysis by an electron microscope, so that particles with the sizes of about 1 micron and conical shapes can be found on the reticular membrane.
Scanning parameters of an electron microscope: electron acceleration voltage 10kV, working distance 6.2mm, magnification 20000 times (as shown in fig. 1) or 40000 times (as shown in fig. 2).
The above is not relevant and is applicable to the prior art.
While certain specific embodiments of the present invention have been described in detail by way of illustration, it will be understood by those skilled in the art that the foregoing is illustrative only and is not limiting of the scope of the invention, as various modifications or additions may be made to the specific embodiments described and substituted in a similar manner by those skilled in the art without departing from the scope of the invention as defined in the appending claims. It should be understood by those skilled in the art that any modifications, equivalents, improvements and the like made to the above embodiments in accordance with the technical spirit of the present invention are included in the scope of the present invention.
Claims (6)
1. A method for fixing egg vitelline membrane protein is characterized in that: the egg is treated to obtain a yolk membrane, and then the yolk membrane is dyed by a Golgi-Cox reagent and then blackened by LiOH.
2. The method for immobilizing egg vitelline membrane proteins of claim 1, wherein: the specific operation of processing the eggs to obtain the vitelline membrane is as follows: sterilizing whole fresh egg with 75% alcohol, breaking egg shell, pouring egg white and yolk into a beaker, sucking out egg white with a disposable straw, sucking out yolk membrane, sucking out yolk in the yolk membrane, and rinsing the yolk membrane with distilled water.
3. The method for immobilizing egg vitelline membrane proteins according to claim 2, wherein: the rinsed yolk membrane was stained in Golgi-Cox reagent for 2 days and 1% LiOH was blackened for one day.
4. A method of immobilising egg vitelline membrane proteins as claimed in claim 3, wherein: the Golgi-Cox reagent comprises the following components in parts by mass: 1 part of mercuric chloride, 1 part of potassium dichromate, 0.8 part of potassium chromate and 80 parts of distilled water.
5. The method for immobilizing egg vitelline membrane proteins of claim 4, wherein: the preparation process of the Golgi-Cox reagent is as follows: firstly, 1 part of mercuric chloride and 1 part of potassium dichromate are dissolved in 64 parts of distilled water, 0.8 part of potassium chromate is dissolved in 16 parts of distilled water in a brown wide-mouth reagent bottle, the reagent bottle is shaken for more than 12 hours at room temperature, then the mercuric chloride and the potassium dichromate are uniformly mixed, the mercuric chloride and the potassium dichromate are kept overnight at the temperature of more than 4 hours or 4 ℃ by a greenhouse shaking table, and a supernatant is taken after a precipitate is deposited at the bottom of the reagent bottle, and the mixture is used for preparation or kept away from light.
6. A method for observing the appearances of egg yolk membrane and membrane protein thereof is characterized in that: fixing egg yolk membrane proteins by the method according to any one of claims 1 to 5, spreading the fixed yolk membrane on a carbon conductive gel, naturally airing, storing in the dark, and performing electron microscope scanning analysis by metal spraying, so that the appearances of the yolk membrane and the membrane proteins thereof can be observed.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011155531.XA CN112504777A (en) | 2020-10-26 | 2020-10-26 | Method for fixing egg yolk membrane protein and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202011155531.XA CN112504777A (en) | 2020-10-26 | 2020-10-26 | Method for fixing egg yolk membrane protein and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN112504777A true CN112504777A (en) | 2021-03-16 |
Family
ID=74955909
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202011155531.XA Pending CN112504777A (en) | 2020-10-26 | 2020-10-26 | Method for fixing egg yolk membrane protein and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN112504777A (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1865901A (en) * | 2006-06-05 | 2006-11-22 | 中国科学院南海海洋研究所 | Seawater fish ovum electron-microscope scanning sample preparing method |
| CN101458182A (en) * | 2008-12-22 | 2009-06-17 | 华中科技大学 | Method for producing brain sample of small animal |
| US20180306688A1 (en) * | 2017-04-21 | 2018-10-25 | Yeu-Kuang Hwu | Method and kit for staining neural tissue sample and method for visualizing neurons |
| US20200284705A1 (en) * | 2017-04-21 | 2020-09-10 | Yeu-Kuang Hwu | Method for visualizing neurons |
-
2020
- 2020-10-26 CN CN202011155531.XA patent/CN112504777A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1865901A (en) * | 2006-06-05 | 2006-11-22 | 中国科学院南海海洋研究所 | Seawater fish ovum electron-microscope scanning sample preparing method |
| CN101458182A (en) * | 2008-12-22 | 2009-06-17 | 华中科技大学 | Method for producing brain sample of small animal |
| US20180306688A1 (en) * | 2017-04-21 | 2018-10-25 | Yeu-Kuang Hwu | Method and kit for staining neural tissue sample and method for visualizing neurons |
| US20200284705A1 (en) * | 2017-04-21 | 2020-09-10 | Yeu-Kuang Hwu | Method for visualizing neurons |
Non-Patent Citations (2)
| Title |
|---|
| BIN ZHANG ET AL.: "Modified Golgi-Cox method for micrometer scale sectioning of the whole mouse brain", 《JOURNAL OF NEUROSCIENCE METHODS》 * |
| MIN AI ET AL.: "Fluorescence imaging of dendritic spines of Golgi-Cox-stained neurons using brightening background", 《J. OF BIOMEDICAL OPTICS》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Bird et al. | The tylenchid (Nematoda) egg shell: structure, composition and permeability | |
| Keil | Contacts of pore tubules and sensory dendrites in antennal chemosensilla of a silkmoth: demonstration of a possible pathway for olfactory molecules | |
| Obokata et al. | Reproducible subcutaneous transplantation of cell sheets into recipient mice | |
| CN104374626A (en) | Moss scanning electron microscope observation material and preparation method thereof | |
| CN108578771B (en) | Preparation method and products thereof with the FGF1 sericin gel for promoting cell-proliferation activity | |
| CN111066736B (en) | Termite embryo development research method | |
| De Santis et al. | A fucosyl glycoprotein component with sperm receptor and sperm-activating activities from the vitelline coat of Ciona intestinalis eggs | |
| CN110779785A (en) | Novel formaldehyde-free tissue fixing solution | |
| CN112504777A (en) | Method for fixing egg yolk membrane protein and application thereof | |
| Leushner et al. | Isolation and characterization of the cuticle from the free-living nematode Panagrellus silusiae | |
| RU2499242C2 (en) | Method for preparing biological tissue samples for histopathological examinations | |
| RU2484446C1 (en) | Method to prepare samples of microorganisms biofilms for survey in scanning electronic microscope | |
| Angel et al. | Ultrastructural patterns of secretory activity in poison cutaneous glands of larval and juvenile Dendrobates auratus (Amphibia, Anura) | |
| Terreni et al. | Development of serous cutaneous glands in Scinax nasica (Anura, Hylidae): patterns of poison biosynthesis and maturation in comparison with larval glands in specimens of other families | |
| CN108543112A (en) | Preparation method and products thereof with the silk gum-agarose plural gel for promoting cell-proliferation activity | |
| Toprak | Light and scanning microscopic structure of filiform papillae in mice. | |
| Bavestrello et al. | Siliceous particles incorporation in Chondrosia reniformis (Porifera, Demospongiae) | |
| Sonpho et al. | ECM-body: A cell-free 3D biomimetic scaffold derived from intact planarian body | |
| Golding | The infracerebral gland in Nephtys—a possible neuroendocrine complex | |
| Sokol et al. | The efficiency of decellularization of bovine pericardium by different concentrations of sodium dodecyl sulfate | |
| WO2022270569A1 (en) | Decellularized cell structure | |
| Rost-Roszkowska et al. | Ultrastructure of the midgut epithelium in Dactylobiotus dispar (Tardigrada: Eutardigrada) during encystation | |
| Pallela et al. | Comparative ultrastructural and biochemical studies of four demosponges from Gulf of Mannar, India | |
| CN113354839B (en) | Collagen-silver nanoparticle composite gel, preparation method and application | |
| Lai-Fook et al. | The importance of methods of fixation in the study of insect blood cells |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210316 |
|
| RJ01 | Rejection of invention patent application after publication |