Technical method of efficiently culturing down producing goat hair follicle stem cell in vitro

A hair follicle stem cell and stem cell technology, which is applied in the field of biomedical stem cells and tissue engineering, can solve problems such as hindering the in-depth development of research and lack of separation and culture methods for hair follicle stem cells, and achieves easy operation and good repeatability.

Active Publication Date: 2013-10-02
QINGDAO AGRI UNIV
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Problems solved by technology

[0005] At present, the in-depth development of research in this field is hindered by the lack of e

Method used

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  • Technical method of efficiently culturing down producing goat hair follicle stem cell in vitro
  • Technical method of efficiently culturing down producing goat hair follicle stem cell in vitro
  • Technical method of efficiently culturing down producing goat hair follicle stem cell in vitro

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Embodiment 1

[0018] 1. Isolation of cashmere goat hair follicles

[0019] Hair follicle stem cells (HFSC) were derived from the dorsal and neck skin of adult cashmere goats. Cut the back and neck skin of cashmere goats, cut off the outer hair to the root, cut into small pieces of 2 square centimeters, and remove the hair follicles from the skin with surgical tweezers, see figure 1 , washed three times in normal saline, soaked in 75% alcohol for 5 minutes, transferred to the operating solution of phosphate buffered saline (PBS, pH 7.0) + 10% fetal calf serum (FCS, Hyclone Company), in the operating solution Wash 3 times, digest with collagenase IV (Sigma) at 37°C for 10 minutes, then separate the hair follicle structure with surgical tweezers under a stereoscope, transfer it to the aforementioned operation solution, and remove the hair follicles connected with it. The sebaceous glands, arrector pili muscle, fat and other impurities were washed 3 times in fresh DMEM / F12 (Hyclone Company). ...

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Abstract

The invention relates to a technical method of efficiently culturing down producing goat hair follicle stem cell in vitro. Firstly, separating down producing goat hair follicle: cutting off back and neck skin of the down producing goat; digesting the back and neck skin of the down producing goat by collagenase IV; separating a hair follicle structure and removing of impurities; secondly, after the hair follicle is washed by culture solution, digesting the hair follicle as single-cells in digestive juice, screening the hair follicle by a mesh with 400 meshes and then transferring the hair follicle in hair follicle stem cell culture solution; regarding the hair follicle stem cell cultured on the day that the hair follicle stem cell transferred into hair follicle stem cell culture solution as zero generation; carrying out cell passage once every 3 to 4 days; thirdly, collecting hair follicle stem cell cloning ball of primary culture culturing 3 to 4 days; blowing and beating the primary cloning ball until to one second or one fourth of the primary cloning ball; moving the cell suspension to new culture solution; according to cell density, carrying out cell passage in ratio of 1 to 2 or 1 to 3. According to the method, a lot of hair follicle stem cells can be obtained from limited material sources, the operation is easy, the repeatability is good, and the cultured hair follicle stem cell can be showed as Beta-intigren, oct-4 and is alkaline phosphatas positive, which shows that the hair follicle stem cell has multilineage differentiation potential.

Description

Technical field: [0001] The invention relates to a technical method for efficiently culturing cashmere goat hair follicle stem cells in vitro, and belongs to the technical field of stem cells and tissue engineering in biomedicine. Background technique: [0002] Stem cells (SC) are a kind of pluripotent cells with self-replication ability, which can differentiate into a variety of functional cells under certain conditions. Stem cells are divided into embryonic stem cells (ES cells) and adult stem cells (somatic stem cells) according to their developmental stages. Embryonic stem cells are totipotent in development, but they only exist in the early stages of embryonic development, so their availability and ethical issues limit their further research and application. [0003] Although adult stem cells do not have the advantages of embryonic stem cells in the development of pluripotency, they are easy to obtain, have a wide range of sources, and still differentiate into various ...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12N5/02
Inventor 孙源超陈春雷李兰沈伟
Owner QINGDAO AGRI UNIV
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