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A transmission electron microscope sample processing method of insect antennae

A transmission electron microscope sample and processing method technology, applied in the field of experimental sample processing, can solve the problem of Spon resin being difficult to penetrate, and achieve the effects of reducing the generation of dyeing precipitate, good polymerization, and beneficial to preservation.

Active Publication Date: 2016-04-27
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The embedding agent can penetrate into the sample more fully through gradient infiltration and the infiltration process on the decolorization shaker, especially solving the problem that the viscous Spon resin is not easy to infiltrate the sample; through the preparation of a new embedding agent, the hardness of the embedding agent can be compared with that of the sample. The hardness of the sample is closer, so that the embedding agent and the sample polymerize well
The embedding plate is used as the embedding mold, which is more conducive to the preparation of the sample and subsequent slicing. The sealing treatment during Spurr polymerization solves the problem of oxygen absorption of Spurr resin

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Preparation of fixative: add sucrose to PBS buffer containing paraformaldehyde and glutaraldehyde, store at 4°C, where PBS buffer is phosphate buffer;

[0030] In 0.1mol / LPBS buffer (pH7.4) containing 2% (mass fraction) paraformaldehyde and 2.5% (mass fraction) glutaraldehyde, add 5% (mass fraction) sucrose, store at 4°C, and phosphoric acid The salt buffer is potassium phosphate buffer or sodium phosphate buffer.

[0031] The specific ratio is as follows: 50ml of 0.2mol / LPBS buffer solution (pH7.4), 10ml of 25% (mass fraction) glutaraldehyde, 20ml of 10% (mass fraction) paraformaldehyde, 20ml of ultrapure water prepared by a pure water meter and 5g of sucrose, prepared by stirring and mixing, and stored at 4°C.

Embodiment 2

[0033] Preparation of fixative: add sucrose to PBS buffer containing paraformaldehyde and glutaraldehyde, store at 4°C, where PBS buffer is phosphate buffer;

[0034] In 0.1mol / LPBS buffer (pH7.4) containing 2% (mass fraction) paraformaldehyde and 2.5% (mass fraction) glutaraldehyde, add 5% (mass fraction) sucrose, store at 4°C, and phosphoric acid The salt buffer is potassium phosphate buffer or sodium phosphate buffer.

[0035] After adding 5% (mass fraction) sucrose, add 1% tannic acid and store at 4°C.

[0036] The specific ratio is as follows: 50ml of 0.2mol / LPBS buffer solution (pH7.4), 10ml of 25% (mass fraction) glutaraldehyde, 20ml of 10% (mass fraction) paraformaldehyde, 20ml of ultrapure water prepared by a pure water meter , 5 g of sucrose and 1 g of tannic acid were prepared by stirring and mixing, and stored at 4°C.

Embodiment 3

[0038] A transmission electron microscope sample processing method of insect antennae, consisting of the following steps successively,

[0039] A. Preparation of fixative: add sucrose to PBS buffer containing paraformaldehyde and glutaraldehyde, and store at 4°C, wherein the PBS buffer is phosphate buffer;

[0040] B. Preparation of various embedding agents:

[0041] (1) SPI-Pon TM Preparation of 812 embedding agent: Add SPI-Pon812, DDSA, NMA and DMP-30 into the beaker one by one, stir evenly with a magnetic stirrer, when all four ingredients are added, seal the mouth of the beaker with aluminum foil, and continue to stir About 1 hour, remove the aluminum foil, use a vacuum desiccator to extract the air bubbles in the embedding agent, and let it stand in the desiccator for a while until the air bubbles completely disappear, then transfer the prepared embedding agent into a dry centrifuge tube or sealed In a syringe, sealed in a plastic bag with silica gel desiccant, stored a...

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Abstract

The invention belongs to the field of experimental sample processing technologies, relates to insect antenna sample processing methods and particularly relates to a transmission electron microscope processing method for insect antenna samples. The method sequentially comprises the following steps of: (A) preparing fixing liquid; (B) preparing various embedding agents; (C) dissecting, fixing and rinsing; (D) dewatering and soaking; and (E) gathering, so as to obtain the samples. The method has the advantages that the problems of difficulty in fixing liquid soaking and insufficiency in embedding agent soaking during the process of insect antenna transmission processing are solved, finally-obtained sample slices can be relatively flat, the phenomena of sample wrinkling, damaging and losing are greatly reduced, internal structures of the sample slices are all effectively fixed, and subcellular structures are clearly visible.

Description

technical field [0001] The invention belongs to the technical field of experimental sample processing, and relates to a sample processing method of insect antennae, in particular to a transmission electron microscope sample processing method of insect antennae. Background technique [0002] Commercial transmission electron microscopes have played a major role in the life sciences since their general introduction to the market in the late 1980s. Conventional transmission electron microscopy sample preparation technology is also very mature. However, there are still certain technical bottlenecks when studying biological samples with dense tissue and high degree of ossification, such as insect antennae. [0003] There are a large number of sensory organs distributed on the antennae of insects, which are very important sensory organs of insects, and play a key role in important ecological behaviors such as insects looking for mates and host plants, and avoiding predators. Stud...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/28
Inventor 付丙鲜祝增荣洪健
Owner ZHEJIANG UNIV
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