CN102589954B - Method using transmission electron microscope to observe ultra microstructure of inorganic high polymer material - Google Patents
Method using transmission electron microscope to observe ultra microstructure of inorganic high polymer material Download PDFInfo
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- CN102589954B CN102589954B CN201110008025.2A CN201110008025A CN102589954B CN 102589954 B CN102589954 B CN 102589954B CN 201110008025 A CN201110008025 A CN 201110008025A CN 102589954 B CN102589954 B CN 102589954B
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Abstract
The invention belongs to the technical field of polymer materials, and relates to a method using a transmission electron microscope to observe ultra microstructure of inorganic high polymer material. The method includes following steps: placing the high polymer material into absolute ethanol solution or absolute acetone solution for eight hours for infiltration; then placing the high polymer material into mixed liquid of absolute ethanol and epoxy resin or mixed liquid of absolute acetone solution and epoxy resin for 2 to 7 days; placing the high polymer material into a 37 DEG C to 45 DEG C baking oven for 11 to 24 hours, so as to remove gaseous ethanol or gaseous acetone and store the high polymer material in pure epoxy resin; furthermore, adding the required amount of the pure epoxy resin to embedded blocks for 24-hour curing. The method provided by the invention can solve the problem that for a long time the embedded high polymer material can not or is difficult to acquire integral ultrathin sections, and performs the positive function for the three-dimensional reconstruction of the physical models inside the high polymer material particles. The method disclosed by the invention is further applied to study in other fields, and is widely applied in study fields such as medical treatment, space flight, and composition of the high polymer material.
Description
Technical field
The invention belongs to technical field of polymer materials, relate to a kind of Ultrastructural method of transmission electron microscope observation inorganic macromolecule material of using, be specifically related to a kind of method of the ultrastructure of inorganic polymer particulate material being cut into slices with transmission electron microscope.
Background technology
Transmission electron microscope is generally used for biological and organic Ultrastructural observation, the electron microscopic sample preparation of most organic-biological materials is ripe and extensive utilization, and the preparation of the electron microscopic sample of inorganic macromolecule material rests on the structure observation of molecule external model always, cannot arrive by transmission electron microscope observation the ultrastructure of macromolecule material particle inside; And along with macromolecular material is at constantly application development of the field such as medical science, space flight; for the fine structure of macromolecular material and synthetic understanding; there is further demand; seek new research method and operational means; and then more the fine structure of studying macromolecular material is understood on deep layer ground, has very actively profound significance for practical application.
At present, conventionally adopt class negative staining technology to inorganic polymer particulate material as SiO
2particle, the multiprotein complex of C nano particle, C nanotube and other organic material particles carries out Electron microscopic study, this technology is in the time carrying out the Ultrastructural observation of electron microscope to polymeric particles material, the result obtaining is the external morphological structure of polymeric particles material or the particle diameter of particle, cannot observe the structure that obtains grain diameter inside.
Research is found, because inorganic macromolecule material relative density is larger, need not equally with biological organization material be fixed, the processing such as dehydration, but machine macromolecular material is after pure epoxy resin 618 embedding immersions are saturating, support because parts of fine microstructure does not have epoxy resin to connect, in the time carrying out ultra-thin section, be prone to the situation that part-structure is lost.
Summary of the invention
The object of the invention is to overcome defect and the deficiency of prior art, a kind of Ultrastructural method of inorganic macromolecule material of observing is provided, especially use the Ultrastructural method of transmission electron microscope observation inorganic macromolecule material.This method improves the method for conventional transmission electron microscope Bio-specimen Preparation, can utilize the Ultrastructural observation of transmission electron microscope to inorganic polymer particulate material and cut into slices.
The invention provides a kind of embedding method of independently macromolecular material; under the effect of liquid-liquid-gas equilibrium principle; macromolecule material particle or single tube inside the support of epoxy resin with link under; form fixing Polymer Molecular Weight; obtain the space diagram structure of narrow distribution, and can use the inside ultrastructure of the above-mentioned macromolecule material particle of conventional transmission electron microscope observation.
Particularly, the Ultrastructural method of transmission electron microscope observation inorganic macromolecule material of using of the present invention, is characterized in that, it comprises step:
(1) macromolecular material is put into 100% straight alcohol or 100% pure acetone solution, soaked into 8 hours;
(2) macromolecular material is put into ethanol or acetone and epoxy resin mixed liquor 2～7 days;
(3) macromolecular material is put into temperature and is in the baking oven of 37 DEG C～45 DEG C and toast, get rid of ethanol or acetone gas, macromolecular material is kept in pure epoxy resin;
(4) add the expense of pure epoxy resin to embedded block, solidify 24 hours.
In the present invention, inorganic macromolecule material comprises: silicon dioxide (SiO
2), organic glass, CNT, carbon dust (C nano particle, C nanotube) etc.
In the present invention, in the described mixed liquor of step (2), the ratio of ethanol or acetone and epoxy resin is 1
:1 to 1
:4;
In the present invention, the described baking time of step (3) is 11～24 hours, and preferably baking time is 12 hours.
In the present invention, described acetone is the solution inflammable, volatile, chemical property is more active, volatilize, and aggretion type epoxy resin 618 (E-51) is metastable solution under certain temperature conditions with vapor form; According to liquid-liquid-gas equilibrium principle, the intermiscibility solution mixing with aggretion type epoxy resin 618 when acetone, along with the vaporization of acetone, the mixed liquor of acetone and aggretion type epoxy resin 618 is all the time in liquid-liquid-gas balance, aggretion type epoxy resin 618 concentration in mixed liquor progressively improve, until aggretion type epoxy resin 618 purifying, curing.
While using the inventive method to carry out embedding, slicing treatment to macromolecular material, result shows, the mixed liquor of acetone and aggretion type epoxy resin 618 is penetrated into SiO
2in particle, C nano particle, C nanotube or organic glass (polymethylmethacrylate, PMMA) macromolecular material high density granular inner structure, after acetone in acetone and aggretion type epoxy resin 618 mixed liquors vaporization volatilization, macromolecular material is as SiO
2aggretion type epoxy resin 618 concentration in the high density gaps such as particle, C nano particle, C nanotube are also thereupon than increasing, until epoxy resin 618 purifying, curing, cause macromolecule material particle interior detail microstructure to obtain connection and the support of epoxy resin, carry out ultra-thin section, make complete macromolecule material particle interior detail microstructure section; In one embodiment of the invention, organic glass macromolecule transparent material (polymethylmethacrylate, PMMA) is carried out to embedding, sectioning, can obtain complete section, need not vulcanize and just can carry out electron microscopic observation.
Advantage by the Ultrastructural method of transmission electron microscope observation inorganic macromolecule material of the present invention is:
Adopt the embedding of method of the present invention to macromolecular material nano particle electron microscope sample, solve a difficult problem that cannot or be difficult to obtain complete ultra-thin section after macromolecular material embedding for a long time, solve more deep structure research method of macromolecular material.Through the macromolecular material of this method embedding, in slicing processes, the injury tolerance of the expensive diamond cutter edge of a knife is reduced to minimum; In the time carrying out the ultra-thin section of sample, obtain serial section, for the three-dimensional reconstruction of the physical model of macromolecule material particle inside has played positive effect.
Method of the present invention is conducive to set up Three-dimension Reconstruction Model, and Three-dimension Reconstruction Model is as micro-targeting vector in medical treatment, has unlimited meaning for the treatment of particular patient's; The inventive method is also applied to the research in other field, is widely used in the research field such as compound of medical treatment, space industry, macromolecular material.
For the ease of understanding, below by the drawings and specific embodiments, the present invention is described in detail.It needs to be noted, specific embodiments and the drawings are only in order to illustrate, obviously those skilled in the art can, according to explanation herein, carry out various corrections or change to the present invention, within these corrections and change also will be included the scope of the invention in.
Brief description of the drawings
Fig. 1 is that carbon granule drips chip architecture figure, shows the particle diameter and the configuration of surface that adopt drop adsorption method can only obtain macromolecular material.
Fig. 2 is silica dioxide granule structural drawing.
Fig. 3 is the structural drawing that obtains complete nano-carbon powder after embedded section.
Fig. 4 is the fine structure figure of CNT.
Fig. 5 is the structural drawing of the carbon pipe in Double-layer clamp film.
Fig. 6 be can serial section device outside drawing.
Specific embodiment
embodiment 1observe silica dioxide granule ultrastructure
Preparation SiO
2the electron microscope sample of particle macromolecule transparent material particle: grind to form the powder of micron utmost point with agate mortar, collect a small amount of powder and put 37
0c baking oven is baked and banked up with earth the part of anhydrating, and a E618 epoxy resin adds 2 part of 100% ethanol or acetone is made embedding medium mixed liquor; Choosing above-mentioned particulate material is placed in imbedded mold on a small quantity, the embedding medium mixed liquor (embedding medium stoste is less than 100% pure acetone or ethanol) of preparation is measured and added in mould on demand, after stirring with fine needle, carry out soaking into early stage curing, put 45 DEG C of baking ovens 12 hours, pure embedding medium is filled it up with in injection in second day, puts 45 DEG C of baking ovens and continues to solidify for 24 hours, direct electron microscopic examination after section, result demonstration, this method can obtain good SiO
2particle macromolecule transparent material granule interior is as the map architecture in honeybee room (as shown in Figure 2).
embodiment 2observe C nano particle, C nanotube ultrastructure
(2) material is CNT, needs a epoxy resin to add 4 part of 100% acetone and makes embedding medium mixing
Prepare the electron microscope embedding sample of macromolecular material C nano particle, C nanotube: for the electron microscope sample embedding of C nano particle and synthetic C nanotube, because the moment absorption affinity of carbon is easily stopped up the high density passage of macromolecular material C nano particle, C nanotube, so, before to the embedding of macromolecular material C nano particle, C nanotube, will be through 37
0c nano particle after C baking oven is baked and banked up with earth or C nanotube are inserted a small amount of 100% acetone of injection in mould, according to liquid-liquid equilibrium principle, pure 100% acetone is introduced 1 part of epoxy resin and is added 4 part of 100% acetone mixed solution, makes it to reach balance, carries out soaking into early stage purifying, solidifies with fine needle after stirring; Put 45 DEG C of baking ovens 12 hours, pure embedding medium is filled it up with in injection in second day, puts 45 DEG C of baking ovens and continues to solidify for 24 hours, direct electron microscopic examination after section, result shows, can obtain the picture structure (as shown in Figure 3) of good macromolecular material C nano particle, C nanotube inside.
embodiment 3observe organic glass macromolecule transparent material ultrastructure
The chemical name of the organic glass macromolecule transparent material of the present embodiment is polymethylmethacrylate (PMMA), is formed by methyl methacrylate polymerization.Adopt embedding method of the present invention to carry out sample preparation to polymethylmethacrylate (PMMA): first block organic glass to be filed out on file to powder strong, collected a small amount of powder and put 37
0c baking oven is baked and banked up with earth the part of anhydrating, because being dissolved in acetone, polymethylmethacrylate (PMMA) is difficult for being dissolved in ethanol, therefore, embedding medium can not can only dilute with 100% ethanol by acetone diluted, selecting a E618 epoxy resin to add 2 ~ 3 part of 100% ethanol measures on demand and adds in mould, after stirring with fine needle, carry out soaking into early stage, purifying, curing, put 45 DEG C of baking ovens 12 hours, within second day, inject and fill it up with pure embedding medium, put 45 DEG C of baking ovens and continue to solidify for 24 hours, after section, in double dish, exert a gradual, corrupting influence on and within 20-30 minutes, carry out electron microscopic observation with 2% osmic acid.
The present invention confirms through experiment, be conducive to set up Three-dimension Reconstruction Model, and Three-dimension Reconstruction Model is as micro-targeting vector in medical treatment, has unlimited meaning for the treatment of particular patient's; The present invention also can be used for the research in other field, is widely used in the research field such as compound of medical treatment, space industry, macromolecular material.
Claims (3)
1. by the Ultrastructural method of transmission electron microscope observation inorganic macromolecule material, it is characterized in that, it comprises step:
(1) macromolecular material is put into straight alcohol or pure acetone solution, soaked into 8 hours;
(2) macromolecular material is put into the mixed liquor of ethanol and epoxy resin or macromolecular material is put into acetone and epoxy resin mixed liquor 2～7 days;
(3) macromolecular material is put into temperature and is in the baking oven of 37 DEG C～45 DEG C, toast 11～24 hours, get rid of ethanol or acetone gas, macromolecular material is kept in pure epoxy resin;
(4) add the expense of pure epoxy resin to embedded block, solidify 24 hours;
Described inorganic macromolecule material is selected from silicon dioxide, organic glass, CNT or carbon dust.
2. by method claimed in claim 1, it is characterized in that, in described step (2), the ratio of ethanol or acetone and epoxy resin is 1:1 to 1:4.
3. by method claimed in claim 1, it is characterized in that, in described step (3), baking time is 12 hours.
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|---|---|---|---|---|
| WO2003099222A2 (en) * | 2002-05-23 | 2003-12-04 | Invitrogen Corporation | Pseudo-tissues and uses thereof |
| CN1587959A (en) * | 2004-09-23 | 2005-03-02 | 上海交通大学 | Method for producing biolgoical sample embedded block based on atomic force microscope observation |
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| US20040219588A1 (en) * | 2003-04-30 | 2004-11-04 | Masaru Furuta | Method for dispensing reagents onto biological samples and method for analyzing biological samples |
| JP2006058270A (en) * | 2004-08-24 | 2006-03-02 | Sumitomo Chemical Co Ltd | Analytical method for cross section of object to be inspected |
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| WO2003099222A2 (en) * | 2002-05-23 | 2003-12-04 | Invitrogen Corporation | Pseudo-tissues and uses thereof |
| CN1587959A (en) * | 2004-09-23 | 2005-03-02 | 上海交通大学 | Method for producing biolgoical sample embedded block based on atomic force microscope observation |
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