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A method for efficiently isolating and culturing primary hepatocytes of Puffer puffer obscura

A dark-striped pufferfish, primary hepatocyte technology, applied in cell dissociation methods, animal cells, tissue culture and other directions, can solve problems such as difficulty in hepatocyte separation, and achieve the solution of difficult hepatocyte separation, high separation efficiency, and physiological steady state effect

Active Publication Date: 2021-05-11
NANJING NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The accumulation of high-density fat in the liver brings great difficulties to the isolation of hepatocytes

Method used

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  • A method for efficiently isolating and culturing primary hepatocytes of Puffer puffer obscura
  • A method for efficiently isolating and culturing primary hepatocytes of Puffer puffer obscura
  • A method for efficiently isolating and culturing primary hepatocytes of Puffer puffer obscura

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Embodiment 1

[0027] 1. A method for efficiently separating and culturing primary hepatocytes of pufferfish obscura, the specific operation steps are:

[0028] (1) Select puffer obscura body length as 2 ± 0.5cm, body weight as 2 ± 0.3g without feed conversion, juvenile fish fed with red worms or Artemia (raised at a salinity of 1-2 before taking liver tissue). 3 weeks in the water environment, starvation treatment 48h), use 70% alcohol cotton balls to wipe the fish body, aseptic dissection, Hank's solution washes for many times to remove impurities, the obtained liver tissue is as follows: figure 1 shown;

[0029] (2) Put the cleaned liver into the pre-prepared type II collagenase (1mg / mL) digestion solution (the dosage is 8ml per gram of liver tissue), and cut the liver into pieces with high-temperature sterilized ophthalmic scissors about 1-3mm 3 size. Transfer the liver fragments together with the collagenase II digestion solution to a 50ml sterile centrifuge tube, and digest them on ...

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Abstract

The invention discloses a method for isolating and culturing primary hepatocytes of puffer puffer, which comprises the following steps: (1) selecting juvenile puffer puffer, aseptically dissecting the liver, cleaning and cutting into pieces; The shredded liver is digested with collagenase II digestion solution, filtered, centrifuged, and the lysate is added to the precipitate to obtain a cell suspension, which is then added to the separation solution and centrifuged to remove impurities; (3) Add pre-warmed Mix the complete medium, make up the complete medium, and culture. Regarding the prior art, the method for separating and culturing primary hepatocytes of Fugu obscura of the present invention can solve the problem of difficulty in separating hepatocytes, and the separation efficiency of hepatocytes is high. training requirements.

Description

technical field [0001] The invention relates to a high-efficiency separation and culture method for primary hepatocytes of puffer puffer, and belongs to the technical field of separation and cultivation of primary liver cells of puffer puffer. Background technique [0002] Fugu (Takifugu fasciatus) is a migratory fish in sea and freshwater, mainly distributed in the middle and lower reaches of the Yangtze River, and is one of the three delicacies of the Yangtze River. In recent years, the state and the government have increased their support for the puffer fish farming industry. The farming technology of puffer puffer obscura has matured, and the output has increased year by year. However, with the rapid development of the industry, various problems in the breeding process have gradually emerged (such as the degradation of germplasm resources and the frequent occurrence of various diseases). Therefore, increasing the basic research on Puffer obscura is conducive to the heal...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
CPCC12N5/067C12N2509/00
Inventor 尹绍武魏小珍王涛胡亚东孙亦如
Owner NANJING NORMAL UNIVERSITY
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