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Method for establishing lipopolysaccharide (LPS)-induced milk cow mammary gland epithelial cell apoptosis model

A technique for mammary epithelial cells and a method for establishing a method is applied in the field of establishing an apoptosis model of lipopolysaccharide-induced mammary epithelial cells in dairy cows, and can solve the problems of little understanding and the like

Inactive Publication Date: 2015-07-08
ANHUI AGRICULTURAL UNIVERSITY
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Problems solved by technology

The latest research results show that mammary gland epithelial cells may have active coping responses in the process of mammary gland infection, but the mechanism is still poorly understood, and it is currently a research hotspot at home and abroad

Method used

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  • Method for establishing lipopolysaccharide (LPS)-induced milk cow mammary gland epithelial cell apoptosis model
  • Method for establishing lipopolysaccharide (LPS)-induced milk cow mammary gland epithelial cell apoptosis model
  • Method for establishing lipopolysaccharide (LPS)-induced milk cow mammary gland epithelial cell apoptosis model

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Embodiment 1

[0042] A method for establishing an apoptosis model of lipopolysaccharide-induced milk bovine mammary epithelial cells, comprising the following steps:

[0043] (1) Cultivate cow mammary epithelial cells, and obtain 2-3 generations of logarithmic growth phase cells for subsequent experiments;

[0044] (2) Take the 2-3 generations of logarithmic growth phase dairy cow mammary epithelial cells, and use 1×10 4 The density of cells / well was inoculated into 96-well plates, and randomly divided into 6 test groups. The cell inhibition rate of the 6 test groups was detected by the CCK-8 method; the setting conditions of the 6 test groups were: to the 100 μL of culture medium containing LPS with different mass concentrations were added to each well of the 96-well plate of the six experimental groups, and the different mass concentrations were 0, 10, 20, 50, 100 and 200 μg / mL, respectively. The action time periods are set to include 3, 8, 16 and 24h respectively; each group / each time p...

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Abstract

The invention discloses a method for establishing a lipopolysaccharide (LPS)-induced milk cow mammary gland epithelial cell apoptosis model, which comprises the following steps: (1) acquiring 2nd-3rd-generation milk cow mammary gland epithelial cells; (2) detecting inhibiting action of LPS on milk cow mammary gland epithelial cell proliferation by a CCK-8 process, and primarily detecting the LPS action concentration and action time which conform to the requirement that the apoptosis rate of the established in-vitro cell trauma model is 40-50%; (3) detecting the mammary gland epithelial cell apoptosis rate by flow cytometry; (4) detecting variation conditions of apoptosis cell nucleic acids by DNA ladder; and (5) observing the interior morphological structure variation of the mammary gland epithelial cells by a transmission electron microscope. The determination method of the LPS-induced milk cow mammary gland epithelial cell apoptosis model induction conditions has the advantages of ideal effect and low cost, and is convenient to operate.

Description

technical field [0001] The invention relates to a method for establishing an in vitro cell apoptosis model, in particular to a method for establishing an apoptosis model of dairy cow mammary epithelial cells induced by lipopolysaccharide. Background technique [0002] Mastitis often occurs in ruminants such as dairy cows and dairy goats, and is an inflammatory reaction of the mammary gland caused by pathogenic bacteria, which can lead to serious economic losses. At present, scholars from all over the world are trying to explore a typical, inexpensive and easy-to-establish animal model of mastitis to study the pathogenesis of mastitis and seek better prevention and treatment drugs. At present, there have been many reports at home and abroad about the production of animal models of mastitis by perfusing pathogenic bacteria and endotoxins into the mammary glands of animals such as cows, goats, and mice. However, due to the high cost of experimental cow and goat mastitis models,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/02
Inventor 丁月云殷宗俊张晓东孟云张威付坤张梦琦黄龙王源朗朱树娇
Owner ANHUI AGRICULTURAL UNIVERSITY
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