Primary culture and purification method of intestinal epithelial cells of carassius auratus gibelio

A technology of intestinal epithelial cells and heterogeneous gibel carp, applied in the field of cultured aquatic biological tissue and cell culture engineering, can solve problems such as research difficulties, affecting the sampling and purification of crucian carp intestinal epithelial cells, and unclear distances between intestinal tissue blocks , to achieve the effect of shortening the incubation time

Inactive Publication Date: 2013-05-29
ZHEJIANG GONGSHANG UNIVERSITY
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Problems solved by technology

The performance of these functions depends on the division, differentiation and growth of intestinal epithelial cells. In fact, due to the differences between fish species, the interaction of many variable factors in the body system and the local microenvironment, the effect on fish intestinal epithelial cells There are still great difficulties in related research, and it is necessary to conduct separate studies on different species of fish
[0005] Primary culture of crucian carp intestinal epithelial cells has been report

Method used

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  • Primary culture and purification method of intestinal epithelial cells of carassius auratus gibelio
  • Primary culture and purification method of intestinal epithelial cells of carassius auratus gibelio

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Embodiment 1

[0017] A method for isolating, purifying, culturing and subcultureing intestinal epithelial cells of heterogeneous gibel crucian carp, comprising the following steps:

[0018] (1) Sampling of intestinal epithelial tissue: select 3 healthy heterogeneous gibel crucian carp (64±5g), fast for 24 hours, anesthetize (MS-222, Sigma; 1:2500), put them in 70% ethanol for 1 min for disinfection. Take out the middle intestinal tract in a sterile room, cut off the mesentery and fat, cut the intestinal wall longitudinally, wash it with PBS buffer several times, remove the intestinal mucus, and cut the intestinal wall to a volume of 0.9-1.1mm 3 organization block, spare.

[0019] (2) Preparation of culture medium: The culture medium was obtained according to the following feed ratio, and 0.1 μg of epithelial cell growth factor, 0.1 IU of insulin, 100 IU of penicillin, 100 μg of streptomycin and 0.1 ml of fetal bovine serum were added to each ml of DMEM medium.

[0020] (3) Tissue block cul...

Embodiment 2

[0024] A method for isolating, purifying, culturing and subcultureing intestinal epithelial cells of heterogeneous gibel crucian carp, comprising the following steps:

[0025] (1) Sampling of intestinal epithelial tissue: select 3 healthy heterogeneous gibel crucian carp (75±4g), anesthetized (MS-222, Sigma; 1:2500) after fasting for 24 hours, and disinfected in 70% ethanol for 1 min. Take out the middle intestinal tract in a sterile room, cut off the mesentery and fat, cut the intestinal wall longitudinally, wash it with PBS buffer several times, remove the intestinal mucus, and cut the intestinal wall to a volume of 0.9-1.1mm 3 organization block, spare.

[0026] (2) Preparation of culture medium: Add 0.1 μg epithelial cell growth factor, 0.1 IU insulin, 100 IU penicillin, 100 μg streptomycin and 0.1 ml fetal bovine serum to each ml of DMEM (Dulbecco’s Modified Eagle’s Medium) medium.

[0027] (3) Tissue block culture: Add 0.25% trypsin to the heterogeneous gibel carp intes...

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Abstract

The invention discloses a primary culture and purification method of intestinal epithelial cells of carassius auratus gibelio. The method comprises the following steps of: sampling intestinal epithelial tissues, preparing a culture solution, culturing tissue blocks and purifying the intestinal epithelial tissues. According to the method disclosed by the invention, processes of separating, culturing and purifying the intestinal epithelial cells of the carassius auratus gibelio, and ingredients of the culture solution, are improved. According to the primary culture method of the intestinal epithelial cells of the carassius auratus gibelio, the time for culturing to normal cohesion of the cells can be shortened to 4-8 hours, the culture time is greatly shortened, and a constructed intestinal epithelial cell model of carassius auratus gibelio can be used for carrying out related researches on nutrient digestion and absorption mechanisms, the cell toxicity of foreign substances and the development and metabolism of the cells.

Description

technical field [0001] The invention relates to the field of cultured aquatic biological tissue and cell culture engineering, in particular to an improved primary culture method for heterogeneous gibel crucian carp intestinal epithelial cells. Background technique [0002] Cell culture is to separate cells from organisms, simulate the physiological environment in vivo in vitro, and make them survive and grow in an artificial environment, so as to study the structure, function, metabolism of cells and the influence of cells on environmental factors more directly. reaction etc. At present, cell culture is a rapidly developing experimental technique in modern biological sciences, which has made important contributions to the research and application of cytology, genetics, virology, and immunology. [0003] The culture technology of animal cells started in 1907 when Harrison successfully cultured the neural plates of frogs in the lymphatic block. It is much more difficult due ...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 王彦波傅玲琳
Owner ZHEJIANG GONGSHANG UNIVERSITY
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