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Method for cleaning and decomposing waxy fixed animal tissue

A technology for fixing animals and wax, which is applied to the extraction and purification of nucleic acid substances in paraffin-fixed tissue sections, and cleanly decomposes waxy fixed animal tissues. It can solve the problems of nucleic acid fragmentation, complicated experimental process, and strong tissue damage. Achieve the effect of quality and quantity guarantee, safe experimental operation and simple operation

Active Publication Date: 2019-03-29
杭州倍强医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The method of xylene and absolute ethanol dissolves paraffin most thoroughly, but xylene is volatile, has a heavy odor, is highly toxic, and is highly destructive to tissues, so it is not a perfect method
The half-hour boiling treatment not only makes the experimental process complicated, but also has certain risks.
Especially under the strict requirements of the current laboratory fire safety, boiling treatment has many inconveniences
At the same time, the destructive effect of NaOH lye and high-temperature boiling make the already fragile and fragile nucleic acid in paraffin tissue more fragmented, affecting the quality of nucleic acid

Method used

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  • Method for cleaning and decomposing waxy fixed animal tissue

Examples

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Comparison scheme
Effect test

Embodiment 1

[0041] The following is a case of implementing the method of the present invention for a moderate amount of paraffin tissue section:

[0042] as attached figure 1 In the description, the 1.5ml plastic centrifuge tube 01 contains 10 coiled and columnar lung cancer paraffin-embedded tissue sections 02 with a thickness of 6 microns and an area of ​​about 4cm×3cm. Inject 250 microliters of dewaxing solution 04 into centrifuge tube 01. The dewaxing solution 04 contains 2% SDS by mass, Tris with a molar concentration of 80mM, EDTA with a molar concentration of 2mM, and the pH value of the solution is 8.0. After injecting dewaxing solution 04, the centrifuge tube was centrifuged at 11,000 g for 1 minute. After centrifugation, the paraffin-embedded tissue sections 02 in the tube are gathered in the lower part of the centrifuge tube under the centrifugal force to form a shrinkage paraffin tissue 03, and the injected dewaxing solution 04 is located above the shrinkage paraffin tissue ...

Embodiment 2

[0044] The following is a small amount of paraffin tissue sections adopting the embodiment case of the method of the present invention:

[0045] as attached figure 1 In the description, centrifuge tube 01 is equipped with a colon cancer paraffin-embedded tissue section 2 with a thickness of 4 microns and a size of 1 square centimeter. Inject 100 microliters of dewaxing solution 04 into centrifuge tube 01. The dewaxing solution 04 contains 0.2% SDS by mass, 1 mM EDTA molarity, 50 mM Tris molarity, and a pH of 7.0. Centrifuge at a high speed of 8000 g for 30 seconds. It can be seen that the paraffin tissue 03 is compressed at the bottom of the centrifuge tube 01 and immersed in the dewaxing solution 04. Place the centrifuge tube 01 in a 60°C water bath and incubate for 5 minutes. Immediately after taking it out, centrifuge at a high speed of 8,000 g for 30 seconds. It can be seen that the wax in the centrifuge tube 01 has been completely dissolved from the paraffin-embedded ti...

Embodiment 3

[0047] The following is a large amount of paraffin tissue sections adopting the embodiment case of the method of the present invention:

[0048] as attached figure 1 In the description, the 15ml centrifuge tube 01 contains 50 paraffin-embedded tissue sections of breast cancer 02 with a thickness of 10 microns and a size of 6cm×6cm. Inject 5 ml of dewaxing solution 04 into centrifuge tube 01. Dewaxing solution 04 contains 10% SDS by mass, 10 mM EDTA molarity, 100 mM Tris molarity, and 8.5 solution pH. The centrifuge tube 01 was centrifuged at a high speed of 15,000 g for 5 minutes. It can be seen that the paraffin-embedded tissue section 02 was compressed at the bottom of the centrifuge tube 01 by the centrifugal force and immersed in the dewaxing solution 04. Place the centrifuge tube 01 in a 90°C water bath for 1 hour, take it out and immediately centrifuge at a high speed of 15,000 g for 10 minutes. It can be seen that the wax in the paraffin-embedded tissue has been comp...

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Abstract

The invention discloses a method for cleaning and decomposing a waxy fixed animal tissue. The method comprises processing to a paraffin embedded tissue slice which is accommodated in a centrifugal tube. The operation process comprises the following steps of adding de-waxing liquid and immediately performing high-speed centrifugation; incubating at a middle or high temperature; after taking out ofthe middle or high temperature environment for incubating, immediately performing high-speed centrifugation; injecting protease K solution into the liquid in the centrifugal tube, and incubating at amiddle temperature; performing high-speed centrifugation again, and sucking tissue decomposed solution. According to the method, xylene or boiling water bath is not required, and time for eliminatingthe wax is not required. High tissue decomposition capability is realized. A large amount of tissue can thoroughly deposed for releasing nucleic acid. The method according to the invention has advantages of simple experiment operation, basically no tissue loss, low nucleic acid damage, wide application range, and high suitability for processing ultralow amount to a large amount of paraffin tissueslice.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for cleanly decomposing wax-fixed animal tissues, and is especially suitable for the extraction and purification of nucleic acid substances in paraffin-fixed tissue sections. Background technique [0002] Wax embedding tissue is a common method to preserve animal tissue cell nucleic acid substances, especially when human tissues are preserved in hospitals, paraffin embedding is generally used. When it is necessary to perform nucleic acid molecular detection on preserved tissues, paraffin tissue sections are usually cut for nucleic acid extraction and purification. The key step in extracting nucleic acid substances from paraffin tissue is to separate the tissue from paraffin, completely decompose it and release the intracellular nucleic acid substances into the solution, in order to successfully extract and purify nucleic acids. At present, the most commonly used method for ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/34C12N15/10
CPCC12N15/1003G01N1/34
Inventor 文中伟
Owner 杭州倍强医药科技有限公司
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