CN102944569A - Method for determining microstructure of lipid microsphere/lipid emulsion - Google Patents
Method for determining microstructure of lipid microsphere/lipid emulsion Download PDFInfo
- Publication number
- CN102944569A CN102944569A CN2012105197017A CN201210519701A CN102944569A CN 102944569 A CN102944569 A CN 102944569A CN 2012105197017 A CN2012105197017 A CN 2012105197017A CN 201210519701 A CN201210519701 A CN 201210519701A CN 102944569 A CN102944569 A CN 102944569A
- Authority
- CN
- China
- Prior art keywords
- sample
- micromechanism
- assay method
- fat
- liplid emulsions
- 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
Images
Landscapes
- Sampling And Sample Adjustment (AREA)
Abstract
The invention discloses a method for determining the micromorphology of a lipid microsphere/lipid emulsion with a soft shell structure by use of a scanning electron microscope. The method comprises the following steps of: diluting and dispersing a to-be-tested sample, treating the sample by an anion device so as to positively charge the to-be-tested sample or a sample container carrying the to-be-tested sample, and scanning the microstructure of the surface of the sample by the change of electron beam energy, thus obtaining the good-reproducibility, stable and clear microstructure of the lipid microsphere/lipid emulsion, wherein the spectrogram of a scanning TEM (transmission electron microscope) is adopted in the determination method.
Description
Technical field
The present invention relates to the assay method of a kind of fat microballoon/liplid emulsions micromechanism, be specifically related to a kind of load sample wire netting that adopts in negative ion generating device processing testing sample and the scanning electron microscope thereof, measure the micromorphologic method of the fat microballoon/liplid emulsions with soft shell mechanism.
Background technology
Scanning electron microscope is the modern RESEARCH ON CELL-BIOLOGY instrument of nineteen sixty-five invention, mainly be to utilize the secondary electron signal imaging to observe the configuration of surface of sample, namely remove scanning samples with extremely narrow electron beam, interaction by electron beam and sample produces various effects, wherein mainly is the secondary of sample.Secondary electron can produce the pattern picture that sample surfaces amplifies, and this similarly is to set up chronologically when sample is scanned, and namely uses the method for pointwise imaging to obtain intensified image.Scanning electron microscope is a kind of broad-spectrum multiple function apparatus, has a lot of superior performances, and it can carry out viewing and emulating and analyzing of three-dimensional appearance, carries out the constituent analysis of microcell when observing pattern.Its application is mainly manifested in two aspects: the material morphology observation is analyzed and constituent analysis.Scanning electron microscope is comprised of three parts: vacuum system, electron beam system and imaging system.The scanning electron microscopic observation sample requirement carries out in high vacuum, no matter is water or dehydrating solution, all can produce tempestuously vaporization in high vacuum, not only affects vacuum tightness, contaminated samples, also can destroy the microtexture of sample.Therefore, sample must carry out drying before with electron microscopic observation.
Fat microballoon/liplid emulsions is that medicine is dissolved in the liplid emulsions of making behind water through the phosphatide emulsion dispersion in the fat oil, is a kind of microsome disperse system of sealing take fat oil as soft matrix and by immobilized artificial membrane, and its mean grain size 200nm is called the fat microballoon.Fat microballoon/liplid emulsions is as pharmaceutical carrier, and target is gathered in " targeted therapy " that diseased region has been realized medicine.By scanning electron microscope (TEM), the micromechanism that can observe fat microballoon/liplid emulsions, scanning electron microscope are mainly passed very thin solid sample surface by high-power electron beam, scan the micromechanism of sample surfaces by the variation of beam energy.When using TEM to test, in order to prevent that air intermediate ion and water are for the interference of electron beam, therefore need in vacuum chamber, scan, so just must use the solid sample of white drying, and the form of particulate is more unstable in fat microballoon/liplid emulsions, if concentration is excessive after dry, the effect of the distortion of fat microballoon or polymerization then can occur, so just can't observe the real structure of fat microballoon.And if the lower fat microballoon of working concentration/liplid emulsions solution example because the fat microballoon is made of phosphatide parcel grease, therefore the surface is the hydrophilic radical of phosphatide, is mainly choline (N(CH
4)
3) or amino (NH
2), with positive charge, therefore the high energy electron of meeting attract electrons bundle was black sphere in the Electronic Speculum collection of illustrative plates after these groups dissolved in aqueous solution, white background, and these are different for the common cognition of Electronic Speculum collection of illustrative plates from everybody.In order to address this problem, need to use coloring agent that sample is dyeed.Coloring agent mainly is the group with positive charge, is combined with the phosphatide negative charge group of microsphere surface after the ionization in aqueous solution, makes dried sample with positive charge, the spheroid of background and the white of black will occur like this.But because the polymkeric substance that the TEM test is used or metal-made fine-structure mesh are as sample carrier, polymkeric substance or metal are all with positive charge, the concentration of sample solution itself is rarer with certain electricity repels for the sample particulate, therefore the success ratio of test is very low, reappearance is very poor, all can't observe any fat microballoon in frequent a plurality of samples.
The inventor is by the discovery of concentrating on studies, avoid causing the negative ion generating device of balance weighing drift phenomenon can solve the defective that exists in present available technology adopting scanning electron microscope mensuration fat microballoon/liplid emulsions micromechanism by adopting in the balance weighing, to use, the principle that adopts the electrolysis air to produce ozone ion produces the negative ion air-flow, thus in and the positive charge in sample or the container.Adopt simultaneously negative ion generating device that the load sample container of sample metal-made or polyester is processed rear recycling scanning electron microscope and observe, acquisition that can be stable is the TEM spectrogram of fat microballoon/liplid emulsions micromechanism clearly.
Summary of the invention
The purpose of this invention is to provide a kind of by after test sample and load sample container are processed with negative ion generating device, the method for the fat microballoon that adopts scanning electron microscope to measure to have soft structures/liplid emulsions micromechanism.That the method has overcome in the prior art is low to the success ratio that exists in fat microballoon/liplid emulsions micromechanism, poor reproducibility, to sample destroy serious, amount of samples is large, the defective that can not truly not reflect the sample micromechanism.
The fat microballoon provided by the invention/assay method of liplid emulsions micromechanism comprises testing sample is become certain concentration with diluted, with coloring agent dyeing, measure by scanning electron microscope behind the sample after the employing negative ion generating device processing dyeing and the load sample container of scanning electron microscope after dry.
Wherein, the dilution of testing sample is water.
Wherein, the dilute concentration of sample is 500 ~ 5000 times.
Wherein, coloring agent is phosphomolybdic acid or phosphotungstic acid or acetic acid uranium.
Wherein, negative ion generating device is that the Japanese Shimadzu Stablo of company removes electrical equipment or the German Mei Tele ANTIST-KIT-UN of company goes electrostatic equipment or the YSTP01 of Sartorius AG to remove the static pen.
Wherein, the drying mode of sample is vacuum decompression drying or infrared lamp drying or natural air drying.
Description of drawings
Fig. 1 is the electromicroscopic photograph of comparative example 1.
Fig. 2 is the electromicroscopic photograph of comparative example 2.
Fig. 3 is the electromicroscopic photograph of comparative example 3.
Fig. 4 is the electromicroscopic photograph of comparative example 4.
Fig. 5 is that the electricity of embodiment 1 is competed photo.
Fig. 6 is the electromicroscopic photograph of embodiment 2.
Fig. 7 is the electromicroscopic photograph of embodiment 3.
Embodiment
The embodiment of form is described in further detail content of the present invention by the following examples.But this should be interpreted as that scope of the present invention only limits to following examples.All technology that realizes based on content of the present invention all belong to scope of the present invention.Obviously, according to content of the present invention, according to ordinary skill knowledge and the customary means of this area, under the prerequisite that does not break away from basic fundamental thought of the present invention, can also make modification, replacement or the change of other various ways.
Comparative example 1
Sample: the Alprostadil liposome microsphere preparation electromicroscopic photograph of not processing through staining counter, approximately 500 times of extension rates, the result is as shown in Figure 1.
As shown in Figure 1: in the situation that process without negative staining, the microballoon in the sample is black, and condenses together, and the microballoon quantity that can observe in the visual field is also fewer.
Comparative example 2
Sample: Alprostadil liposome microsphere preparation electromicroscopic photograph under the less extension rate after staining counter is processed, approximately 500 times of extension rates, the result is as shown in Figure 2.
The polymerism that can be observed the fat microballoon by Fig. 2 is apparent in view, is beyond recognition its micromechanism.
Comparative example 3
Sample: Alprostadil liposome microsphere preparation electromicroscopic photograph under the larger extension rate after staining counter is processed, approximately 1000 times of extension rates, the result is as shown in Figure 3.
Can observe the polymerization of fat microballoon by Fig. 3 less, but part distortion is arranged, with carry net in conjunction with relatively poor, behind the replication, poor reproducibility.
Comparative example 4
Sample: the Alprostadil liposome microsphere preparation electromicroscopic photograph that larger extension rate obtained by negative ion generating device after staining counter was processed, approximately 2500 times of extension rates, the result is as shown in Figure 4.
Can observe the polymerization of fat microballoon by Fig. 4 less, deformation is less, and can observe the part micromechanism, and behind the replication, picture is consistent, favorable reproducibility.
Embodiment 1
Sample: alprostadil injection (when trade name is triumphant), the negative staining agent: phosphomolybdic acid, extension rate: approximately 2500 times, negative ion generating device is that the Japanese Shimadzu Stablo of company removes electrical equipment, and drying mode is that vacuum decompression is dry, and test result as shown in Figure 5.
Embodiment 2
Sample: Flurbiprofen ester injection (trade name Furbiprofen axetil), the negative staining agent: phosphotungstic acid, extension rate: approximately 1000 times, negative ion generating device is that the German Mei Tele ANTIST-KIT-UN of company removes electrostatic equipment, drying mode is that vacuum decompression is dry, and test result as shown in Figure 6.
Embodiment 3
Sample: fat emulsion injection (trade name Intralipid), the negative staining agent: acetic acid uranium, extension rate: approximately 4000 times, negative ion generating device is that YSTP01 removes the static pen, and drying mode is that infrared lamp is dry, and test result is as shown in Figure 7.
By Fig. 5,6,7 as can be known, in 3 parts of commercially available samples that adopt the inventive method to measure, the particulate rounding of fat microballoon/liplid emulsions, clear, separation property good, and microscopic pattern is clear, not polymerization adhesion, the good reproducibility of repeatedly measuring.
Claims (6)
1. the assay method of fat microballoon/liplid emulsions micromechanism, it is characterized in that testing sample is become certain concentration with diluted, with coloring agent dyeing, measure by scanning electron microscope behind the sample after the employing negative ion generating device processing dyeing and the load sample container of scanning electron microscope after dry.
2. the assay method of fat microballoon according to claim 1/liplid emulsions micromechanism, the dilution that it is characterized in that testing sample is water.
3. the assay method of fat microballoon according to claim 1/liplid emulsions micromechanism, the dilute concentration that it is characterized in that sample be 500-5000 doubly.
4. the assay method of fat microballoon according to claim 1/liplid emulsions micromechanism is characterized in that coloring agent is phosphomolybdic acid, phosphotungstic acid, acetic acid uranium.
5. the assay method of fat microballoon according to claim 1/liplid emulsions micromechanism is characterized in that negative ion generating device is that the Japanese Shimadzu Stablo of company removes electrical equipment or the German Mei Tele ANTIST-KIT-UN of company goes electrostatic equipment or the YSTP01 of Sartorius AG to remove the static pen.
6. the assay method of fat microballoon according to claim 1/liplid emulsions micromechanism, the drying mode that it is characterized in that sample is vacuum decompression drying or infrared lamp drying or natural air drying.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012105197017A CN102944569A (en) | 2012-12-07 | 2012-12-07 | Method for determining microstructure of lipid microsphere/lipid emulsion |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012105197017A CN102944569A (en) | 2012-12-07 | 2012-12-07 | Method for determining microstructure of lipid microsphere/lipid emulsion |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN102944569A true CN102944569A (en) | 2013-02-27 |
Family
ID=47727531
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2012105197017A Pending CN102944569A (en) | 2012-12-07 | 2012-12-07 | Method for determining microstructure of lipid microsphere/lipid emulsion |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102944569A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104020182A (en) * | 2014-05-27 | 2014-09-03 | 中山大学附属第三医院 | Method for determination of stereo shape and distribution of protein and polypeptide drug loaded microsphere |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1587978A (en) * | 2004-09-24 | 2005-03-02 | 北京工业大学 | Oxygen environment scan electronic microscopic method and system |
| CN1876177A (en) * | 2005-06-06 | 2006-12-13 | 北京圣医耀科技发展有限责任公司 | Biodegradable material microsphere blood vessel suppository containing liposome cell factor and its preparation and uses |
| CN1895223A (en) * | 2006-04-05 | 2007-01-17 | 沈阳药科大学 | Production of elaioplast |
| US20070187597A1 (en) * | 2006-02-13 | 2007-08-16 | Hitachi High-Technologies Corporation | Focused ion beam system and a method of sample preparation and observation |
| JP4185604B2 (en) * | 1998-11-18 | 2008-11-26 | 株式会社日立製作所 | Sample analysis method, sample preparation method and apparatus therefor |
-
2012
- 2012-12-07 CN CN2012105197017A patent/CN102944569A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP4185604B2 (en) * | 1998-11-18 | 2008-11-26 | 株式会社日立製作所 | Sample analysis method, sample preparation method and apparatus therefor |
| CN1587978A (en) * | 2004-09-24 | 2005-03-02 | 北京工业大学 | Oxygen environment scan electronic microscopic method and system |
| CN1876177A (en) * | 2005-06-06 | 2006-12-13 | 北京圣医耀科技发展有限责任公司 | Biodegradable material microsphere blood vessel suppository containing liposome cell factor and its preparation and uses |
| US20070187597A1 (en) * | 2006-02-13 | 2007-08-16 | Hitachi High-Technologies Corporation | Focused ion beam system and a method of sample preparation and observation |
| CN1895223A (en) * | 2006-04-05 | 2007-01-17 | 沈阳药科大学 | Production of elaioplast |
Non-Patent Citations (1)
| Title |
|---|
| 朱莉: "电镜单颗粒技术研究全长AMPK蛋白的构架及变构效应", 《中国博士学位论文全文数据库 基础科学辑》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104020182A (en) * | 2014-05-27 | 2014-09-03 | 中山大学附属第三医院 | Method for determination of stereo shape and distribution of protein and polypeptide drug loaded microsphere |
| CN104020182B (en) * | 2014-05-27 | 2016-04-13 | 中山大学附属第三医院 | The assay method of load albumen, polypeptide drug microballoon solid shape and distribution |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Noble et al. | Direct comparison of optical and electron microscopy methods for structural characterization of extracellular vesicles | |
| Li et al. | The effects of chemical fixation on the cellular nanostructure | |
| Horstmann et al. | Serial section scanning electron microscopy (S3EM) on silicon wafers for ultra-structural volume imaging of cells and tissues | |
| Morris | Modern microscopic methods of bioaerosol analysis | |
| Jusman et al. | Investigation of CPD and HMDS sample preparation techniques for cervical cells in developing computer‐aided screening system based on FE‐SEM/EDX | |
| Miyazaki et al. | Application of low-vacuum scanning electron microscopy for renal biopsy specimens | |
| Lewczuk et al. | Field-emission scanning electron microscope as a tool for large-area and large-volume ultrastructural studies | |
| CN102944569A (en) | Method for determining microstructure of lipid microsphere/lipid emulsion | |
| US9086343B2 (en) | Methods for observing samples and preprocessing thereof | |
| CN106093091A (en) | A kind of rupture of membranes prepares the method for the unicellular scanning electron microscope example of protozoacide | |
| Zhao et al. | The application and development of electron microscopy for three-dimensional reconstruction in life science: a review | |
| CN108693002A (en) | A kind of imitated biological tissue's thin slice, preparation method and its application and device | |
| Lakshman | Diagnostic Electron Microscopy (EM) for Avian Diseases-An Overview | |
| CN105044133B (en) | A kind of simple detection method of transmission electron microscope for high molecule nano material | |
| CN109765202B (en) | Method for rapidly detecting bacterial endotoxin | |
| Weiss et al. | Capillary electrophoresis, gas-phase electrophoretic mobility molecular analysis, and electron microscopy: effective tools for quality assessment and basic rhinovirus research | |
| CN109765164A (en) | A kind of absolute quantitation detection method of unicellular middle gold nanorods | |
| CN113295719A (en) | Method for manufacturing exosome transmission electron microscope sample | |
| Chatterjee et al. | Electron microscopy in the perspective of modern biology: Ultravision and ultradimension | |
| Zhang et al. | Application of electron microscopes in nanotoxicity assessment | |
| Sathananthan et al. | Digital imaging of stem cells by electron microscopy | |
| Zhang et al. | Dictydiaethalium dictyosporangium sp. nov. from China | |
| Newcomb | Modern approaches to the preparation of TEM samples | |
| CN105910999B (en) | It is a kind of to measure devices and methods therefor of the charged single drop to fine particle adsorbance | |
| Chandarana et al. | Classical Wet Mount Method for Observing Live Protists |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20130227 |