Separation and culture method of human skin keratinization layer cells

A technology of skin keratin and culture method, which is applied in the field of cell culture of modern biotechnology, can solve the problems of small quantity, low cell purity, long culture time, etc., and achieve the effect of simple production, good repeatability and easy storage

Inactive Publication Date: 2019-10-29
JIANGYIN CHI SCI
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0004] The cells isolated by the tissue block adherence method are not high in purity, the number is small, and the culture time is long

Method used

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  • Separation and culture method of human skin keratinization layer cells
  • Separation and culture method of human skin keratinization layer cells

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Embodiment Construction

[0022] The terms used in the present invention, unless otherwise specified, generally have the meanings commonly understood by those skilled in the art.

[0023] The present invention will be described in detail below in conjunction with the accompanying drawings and examples, and the protection content of the present invention is not limited to the following examples.

[0024] In the following examples, various procedures and methods not described in detail are conventional methods well known in the art.

[0025] The experimental instruments and reagents used in this experiment are as follows: a set of surgical instruments, CCK-8 cell counting box (Sigma, USA), inverted microscope (XDS-1A, Shanghai), fluorescence microscope (Leica, USA), cryogenic centrifuge ( TD24B-WS, Shanghai), ultra-low temperature refrigerator (Zhongke Meiling), pipette gun (Eppendorf, USA), electronic analytical balance (Sartorius, USA), ultra-clean bench (HJ-CJ-1D, Shanghai), CO 2 Cell culture box (Sa...

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Abstract

The invention provides a separation and culture method of human skin keratinization layer cells. The separation and culture method comprises the following steps of (1) cutting and obtaining fresh skintissue, and transporting the obtained fresh skin tissue back to a laboratory at 4 DEG C; (2) performing cleaning with PBS for 3 times; (3) cutting up the cleaned tissue, and performing digestion withneutral protease at 4 DEG C; (4) separating derma from cuticle; (5) continuing to digest cuticle tissue with preheated trypsin; (6) finishing digestion, performing centrifuging, and discarding supernatant; (7) detecting cell activity through an trypan blue dyeing experiment; (8) after the cells are counted, performing re-suspending with complete culture mediums, performing inoculation to a coatedculture flask, and placing the culture flask in 5% CO2 environment of 37 DEG C for culture; (9) removing fiber forming cells by a differential wall-adherence method; and (10) performing cell identification. The separation and culture method of human skin keratinization layer cells provided by the invention is simple to operate, the obtained cells are high in quantity and high in survival rate, the separation and culture method of human skin keratinization layer cells is ideal, and reliable cell resources are provided for the experiment.

Description

technical field [0001] The invention belongs to the technical field of cell culture of modern biotechnology, and specifically relates to a method for separating and culturing human skin keratinocytes. Background technique [0002] The skin plays an important role in the body's immunity and endocrine, and is one of the physiological barriers of the human body. Keratinocytes are an important component of epidermal cells. The in vitro culture of human skin keratinocytes provides a new way for the research of skin diseases and skin burn treatment. Human skin keratinocytes grow adherently and are oval in shape. The human skin keratinocytes obtained through primary culture can avoid the influence of long-term passage of cell lines on cell characteristics, and provide reliable information for various experimental researches. [0003] Currently, the commonly used methods for isolating human skin keratinocytes include tissue block attachment method and enzyme digestion method. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12Q1/02
CPCC12N5/0629C12N2500/84C12N2501/33C12N2509/00G01N33/5005
Inventor 不公告发明人
Owner JIANGYIN CHI SCI
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