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A kind of preparation method of insect sample for scanning electron microscope

A scanning electron microscope and insect technology, applied in the field of insect sample preparation, can solve the problems of slow penetration of glutaraldehyde, high degree of image shrinkage, sample deformation, etc., to achieve great practical application value, good insect morphological structure, prevent shrunken effect

Inactive Publication Date: 2017-03-01
BEIJING UNIV OF AGRI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Tissue fixation in electron microscope sample preparation generally uses 2%-3% glutaraldehyde, but sometimes the image still shows a high degree of shrinkage, which may be related to the slow penetration speed of glutaraldehyde. And when there is a protective layer on the surface of the sample, it is easy to have poor fixation in the deep part of the tissue. At the same time, the body fluid content of the insect is high, so the sample is more likely to deform, shrink and collapse, which is not conducive to the observation of the hidden place of the sample.

Method used

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  • A kind of preparation method of insect sample for scanning electron microscope
  • A kind of preparation method of insect sample for scanning electron microscope
  • A kind of preparation method of insect sample for scanning electron microscope

Examples

Experimental program
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Effect test

Embodiment 1

[0038] Embodiment 1 (different from the comparative example in step 2)

[0039] The preparation method steps of scanning electron microscope insect sample of the present invention are as follows:

[0040] 1) Gently brush the surface of the larvae of the peach moth borer with an ear washing ball, and then wash the larvae three times with 0.1M phosphate buffer solution (pH7.2);

[0041] 2) Immerse the larvae in the fixative solution (fixative solution is 3% glutaraldehyde), lightly prick three small holes on each side of the larvae under a dissecting microscope, and transfer to a 1.5ml centrifuge tube, which contains fixative Liquid about 1ml. Place the centrifuge tube in a centrifuge tube rack and heat it in a microwave oven on high heat for 8 seconds. At this time, the centrifuge tube is hot but not hot. Replace the fixative in the centrifuge tube and repeat the heating twice, then fix in a refrigerator at 5°C for 12 hours;

[0042] 3) Wash the larvae 5 times with phosphate...

Embodiment 2

[0053] The difference between this embodiment and embodiment 1 is:

[0054] (2) Immerse the larvae in the fixative solution, which is prepared by adding 12 μl Tween-80 to every 20ml of 2.5% glutaraldehyde, lightly puncture three small holes on each side of the larvae under a dissecting microscope, Transfer to a 1.5ml centrifuge tube with about 1ml of fixative in the centrifuge tube. Place the centrifuge tube in a centrifuge tube rack and heat it in a microwave oven on high heat for 8 seconds. At this time, the centrifuge tube is hot but not hot. Replace the fixative in the centrifuge tube and repeat the heating twice, then fix in the refrigerator at 5°C for 12h.

[0055] All the other steps are the same as in Example 1. It can be seen from the imaging results that the abdominal nodes and internodes of the peach borer larvae have no obvious wrinkles compared with Example 1, and the degree of shrinkage is further reduced.

Embodiment 3

[0057]The preparation method of the peach borer larva scanning electron microscope sample comprises the following steps:

[0058] 1) Gently brush the surface of the larvae of the peach moth borer with an ear washing ball, and then wash the larvae three times with 0.1M phosphate buffer solution (pH7.2);

[0059] 2) Soak the larvae in the fixative solution (the fixative solution is prepared by adding 0.018g NaCl and 12μl Tween-80 to 20ml of 2.5% glutaraldehyde), and lightly prick three larvae on each side of the body under a dissecting microscope small hole, transfer to a 1.5ml centrifuge tube, and there is about 1ml of fixative in the centrifuge tube. Place the centrifuge tube in a centrifuge tube rack and heat it in a microwave oven on high heat for 8 seconds. At this time, the centrifuge tube is hot but not hot. Replace the fixative in the centrifuge tube and repeat the heating twice, then fix in a refrigerator at 5°C for 12 hours;

[0060] 3) Wash the larvae 5 times with p...

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Abstract

The invention provides a preparation method of an insect sample for a scanning electron microscope. The preparation method comprises the following steps of: cleaning insect material, fixing tissues, dewatering, replacing, drying and spraying metal and the like, wherein the tissue fixing comprises the steps of: immersing the insect material in stationary liquid, heating in a microwave oven for a period of time, and respectively fixing for a period of time under room temperature and low temperature. The method provided by the invention is rapid and simple and easy to carry out, a clear scanning electron microscope image of a surface structure of an insect can be obtained, the whole morphological structure of the insect can be better maintained, and the deformation, the shrinkage and the collapse are hardly caused. The preparation method has high practical application value.

Description

technical field [0001] The invention relates to a method for preparing an insect sample for a scanning electron microscope. Background technique [0002] Insects are an important part of the animal kingdom. Scanning electron microscopes are widely used in the study of insect structures: scanning electron microscopes are widely used in insect physiology, insect anatomy, and insect systematics. A large number of relevant literature and practical operations show that the preparation of insect samples directly affects the objectivity and authenticity of the observation site. [0003] Insect samples are generally prepared through steps such as sample cleaning, tissue fixation, dehydration, replacement, drying, and gold spraying. Because the overall insect material is large, and the outer skin of the insect has a certain degree of ossification, it is not easy for the fixative to penetrate the internal organs and tissues of the insect. Whether the ultrastructure of each part of t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/28
Inventor 杨瑞杜艳丽王建立郝敬虹王敬贤王绍辉张琳娜房克凤于同泉郝宏京
Owner BEIJING UNIV OF AGRI
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