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A staining method for observing collagen fibers by polarized light microscope

A dyeing method and collagen fiber technology, which is applied in the field of dyeing collagen fibers observed by a polarized light microscope, can solve the problems of unclear structure of collagen fibers, cumbersome steps, weak coloring of collagen fibers, etc.

Active Publication Date: 2021-07-06
GUANGZHOU WOMEN AND CHILDRENS MEDICAL CENTER
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0002] Collagen fibers are widely distributed in various organs and are the most abundant fibers. There are four types of collagen fibers (I, II, III, and IV) known in the prior art. These four types of collagen fibers are important for research The pathogenesis and evolution of lesions have certain significance. Although immunohistochemical staining and molecular pathology techniques have developed rapidly, the application of special staining methods in pathological diagnosis is still irreplaceable; currently commonly used special staining methods for collagen fibers include Masson staining , VanGieson staining, HE staining, picric acid-Sirius red staining methods, etc. Under the light microscope, Masson staining and Van Gieson staining cannot distinguish different types of collagen fibers, and it is difficult to complete the distribution of various types of collagen fibers on tissue sections. Observation and detection of structure and content; and this kind of staining steps are cumbersome and difficult to operate; application of picric acid-Sirius red staining method combined with polarizing microscope observation of collagen fibers can improve resolution; combined with image analysis technology, different types of The type, distribution, arrangement and content of collagen fibers can be analyzed by positioning and quantitative analysis of collagen fibers, which is convenient for judging the degree of tissue fibrosis. This method has incomparable advantages over ordinary light microscope observation
[0003] In the prior art, conventional picric acid-Sirius red staining is used. Under the polarizing microscope, the collagen fibers are weakly stained and the effect is not good. Moreover, ethanol dehydration is often used after the sections are stained, which easily weakens the background of the yellow staining, resulting in collagen fiber structure. Not clear, affecting the correct observation of the tissue and lesion analysis

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  • A staining method for observing collagen fibers by polarized light microscope
  • A staining method for observing collagen fibers by polarized light microscope
  • A staining method for observing collagen fibers by polarized light microscope

Examples

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Embodiment 1

[0034] Tissues were obtained from 42-day-old normal and chronic fibrotic mouse livers and fixed with 10% formaldehyde.

[0035] Picric acid-Sirius red dyeing solution: add 0.1g of picric acid to 1mL Sirius red dyeing solution, after a 90°C water bath, there is insoluble powder at the bottom of the dyeing solution, take the upper solution for later use.

[0036]Staining method: (1) fix the tissue in 10% formaldehyde solution, routinely dehydrate and embed in wax for pathology; (2) paraffin sections with a thickness of 4 μm, routinely dewax to water to obtain tissue sections; (3) drip on the tissue sections Add picric acid-Sirius red staining solution in a water bath at 90°C for 30 minutes; (4) Add 1% w / v acetic acid aqueous solution dropwise and soak for 2 times, each time for 30 seconds; (5) Wash slightly in running water to remove the surface staining (6) Dry in an oven, and fix with neutral resin; (7) Observe under ordinary light microscope and polarized light microscope.

Embodiment 2

[0038] Tissues were obtained from 42-day-old normal and chronic fibrotic mouse livers and fixed with 10% formaldehyde.

[0039] Picric acid-Sirius red dyeing solution: add 0.1g of picric acid to 1mL Sirius red dyeing solution, after a 90°C water bath, there is insoluble powder at the bottom of the dyeing solution, take the upper solution for later use.

[0040] Staining method: (1) fix the tissue in 10% formaldehyde solution, routinely dehydrate and embed in wax for pathology; (2) paraffin sections with a thickness of 4 μm, routinely dewax to water to obtain tissue sections; (3) drip on the tissue sections Add picric acid-Sirius red staining solution in a 70°C water bath for 30 minutes; (4) Add dropwise 1% w / v acetic acid aqueous solution for 2 times, 30s each time; (5) Slightly soak in running water to remove the surface staining of tissue sections (6) Dry in an oven, and fix with neutral resin; (7) Observe under ordinary light microscope and polarized light microscope.

Embodiment 3

[0042] Tissues were obtained from 42-day-old normal and chronic fibrotic mouse livers and fixed with 10% formaldehyde.

[0043] Picric acid-Sirius red dyeing solution: add 0.1g of picric acid to 1ml Sirius red dyeing solution, after 90℃ water bath, there is insoluble powder at the bottom of the dyeing solution, take the upper solution for later use.

[0044] Staining methods: (1) Fix the tissue in 10% formaldehyde solution, routinely dehydrate and make transparent, and embed in wax; (2) Paraffin sections with a thickness of 4 μm are routinely dewaxed to water to obtain tissue sections; (3) Hematoxylin staining for 6 minutes , rinse the tissue sections slightly with running water; (4) stain the tissue sections with picric acid-Sirius red staining solution in a 90°C water bath for 30 min; (5) add 1% w / v acetic acid aqueous solution to soak twice, each time 30s; (6) Slightly immerse in running water to remove the staining solution on the surface of the tissue section; (7) Dry in ...

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Abstract

The invention provides a staining method for observing collagen fibers with a polarized light microscope. The method is to stain with saturated picric acid-Sirius red staining solution at 70°C-90°C, then soak the tissue sections with a weak acid solution with a concentration of 1-10% w / v, and finally soak the tissue sections as usual. Washing, drying, and fixing; the heated picric acid has a stronger penetration effect on the tissue, making the background of the lining more vivid, and the heated Sirius red dye solution is also more firmly adsorbed on the collagen fibers, and the coloring is clear; the weak acid solution can be washed off The crystallization of picric acid precipitated in the tissue section is not easy to wash off the background of the yellow lining, so as to ensure that the collagen fibers are well colored, the background of the yellow lining is obvious, and the collagen fibers are clear; the dyeing method of the present invention has good repeatability, and the collagen fibers are colored Obviously, the fiber structure is clear. This method is conducive to the positioning and quantitative analysis of different types of collagen fibers, and is convenient for judging the degree of tissue fibrosis.

Description

technical field [0001] The invention relates to the technical field of tissue dyeing, and more specifically, to a dyeing method for observing collagen fibers with a polarized light microscope. Background technique [0002] Collagen fibers are widely distributed in various organs and are the most abundant fibers. There are four types of collagen fibers (I, II, III, and IV) known in the prior art. These four types of collagen fibers are important for research The pathogenesis and evolution of lesions have certain significance. Although immunohistochemical staining and molecular pathology techniques have developed rapidly, the application of special staining methods in pathological diagnosis is still irreplaceable; currently commonly used special staining methods for collagen fibers include Masson staining , VanGieson staining, HE staining, picric acid-Sirius red staining methods, etc. Under the light microscope, Masson staining and Van Gieson staining cannot distinguish differ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/30
CPCG01N1/30
Inventor 张锐忠林泽锋付铭童燕陆陈虹交王贺珍陈严夏慧敏
Owner GUANGZHOU WOMEN AND CHILDRENS MEDICAL CENTER
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