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Plant tissue integral staining method for ultrathin slice

A plant tissue and overall dyeing technology, which is applied in the preparation of test samples, sampling, measuring devices, etc., can solve the problems of plant tissue dyeing, solve the problem of lead pollution, and enhance the effect of electronic contrast

Active Publication Date: 2019-11-15
BEIJING FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, plant tissue is different from animal tissue in that it has a cell wall, a feature that poses a huge problem for the overall staining of plant tissue

Method used

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  • Plant tissue integral staining method for ultrathin slice
  • Plant tissue integral staining method for ultrathin slice
  • Plant tissue integral staining method for ultrathin slice

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0022] sample collection

[0023] Collect mature Psychia chinensis pollen and store it at -20°C for later use.

[0024] Sample Preparation

[0025] 1. Pre-fixation: Place the collected mature L. chinensis pollen in a fixative solution composed of 2% glutaraldehyde and 2.4% paraformaldehyde for fixation, and vacuum for 1-2h. Note that the fixative should be stored in a refrigerator at 4°C and placed on ice when in use.

[0026] 2. Rinsing: Rinse the pre-fixed plant samples with 0.1M phosphate buffer solution, wash 3 times, 10-15min each time; store the phosphate buffer solution in a refrigerator at 4°C, and put it on ice when in use.

[0027] 3. Post-fixation: Post-fix the rinsed plant samples with 1% osmic acid, and place them at 4° C. for 2 hours.

[0028] 4. Rinsing: Wash the post-fixed plant samples with pure water for 3 times, each time for 10-15 minutes.

[0029] 5. Prepare 1% TCH: Weigh 0.05g TCH and dissolve it in 5mL pure water, wrap a layer of tin foil to avoid li...

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Abstract

The invention discloses a plant tissue integral staining method for ultrathin slice. The method comprises the following experiment steps of (1) fixing, in which matured picea wilsonii pollen is collected, is pre-fixed by glutaraldehyde and paraformaldehyde and is post-fixed by 1% osmic acid, the matured picea wilsonii pollen is processed by TCH, and osmium is further deposited by 1% osmic acid; (2) block staining, in which block staining is performed with 2.5% gadolinium acetate, 2.5% samarium acetate and 2% uranyl acetate; (3) gradient dehydration with ethyl alcohol, in which dehydration is performed with 30% ethyl alcohol, 50% ethyl alcohol, 60% ethyl alcohol, 70% ethyl alcohol, 80% ethyl alcohol, 90% ethyl alcohol, 95% ethyl alcohol and 100% ethyl alcohol; (4) permeation, in which permeation is performed with an ethyl alcohol and acetone mixed solution, 100% acetone, an acetone and spurr resin mixed solution and 100% spurr resin; and (5) embedding and polymerization, in which 100% spur resin is added into an embedment plate, and a sample is placed and then placed in an oven for polymerization. The method is simple to operate, is relatively good in staining effect and obvious inimage contrast, ultrathin and continuous slicing is also facilitated, and a technical foundation is laid for achieving of plant tissue three-dimensional reconstruction.

Description

technical field [0001] The invention relates to a whole-body dyeing method for plant tissue used for ultrathin sections Background technique [0002] Ultrathin section technique is an important method to study cell submicrostructure. However, in order to obtain images with clear contrast, biological samples must be electronically stained. The so-called electronic staining is to make heavy metals in heavy metal salts such as uranium, lead, osmium, and tungsten combine or be adsorbed with certain components in the tissue to improve the contrast of the fine structure in the sample. There are different numbers of heavy metal atoms adsorbed on different structural components, and the region with more heavy metals (that is, the part with dense structure and high atomic number) has a strong electron scattering ability, which appears as dense black under the electron microscope, and the region with more heavy metals The area with less is light black, gray black, and the area witho...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/28G01N1/30G01N1/36
CPCG01N1/28G01N1/30G01N1/36G01N2001/364
Inventor 李晓娟姚小敏林金星马玲玉沈微微
Owner BEIJING FORESTRY UNIVERSITY
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