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A serum-free culture system for epidermal cell culture

A serum-free medium, epidermal cell technology, applied in artificial cell constructs, animal cells, vertebrate cells, etc., to achieve the effect of large number, low differentiation rate and high cell activity

Active Publication Date: 2019-12-06
NOVAPRINT THERAPEUTICS SUZHOU CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the problems existing in the current serum-containing culture system, the present invention provides a serum-free culture system, which can be applied to culture with trophoblast and without trophoblast at the same time

Method used

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  • A serum-free culture system for epidermal cell culture
  • A serum-free culture system for epidermal cell culture
  • A serum-free culture system for epidermal cell culture

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1, healthy male circumcision post-circumcision skin, in vitro expansion of epidermal cells

[0040] (1) Materials and methods

[0041] Sample: 3cm of circumcised foreskin skin from healthy males discarded after surgery in the hospital 2 size

[0042] Cell culture medium: DMEM, Ham's F12 mixed medium, wherein, DMEM, Ham's F12 weight ratio 3: 1

[0043] Added factors: L glutamine (2mM), cholera toxin (0.1ug / ml), adenine (180uM), hydrocortisone (0.4ug / ml), insulin (20ug / ml), human epidermal growth factor (10ug / ml), transferrin (5ug / ml), triiodothyronine (2uM), CaCl 2 (1 mM).

[0044] (2) Cell isolation and culture

[0045] a) The skin sample is refrigerated and transported to the laboratory in DMEM culture medium, the area is measured, and 3cm is taken 2 size.

[0046] b) Wash the sample with PBS; wash the sample again after disinfection with iodine tincture and alcohol, cut off part of the dermis and chop it into pieces.

[0047] c) 0.05% trypsin cold dig...

example 2

[0052] Example 2, take the breast skin of a healthy adult female after surgery, and expand epidermal cells in vitro

[0053](1) Materials and methods

[0054] Sample: About 3cm of normal breast skin of an adult female discarded during surgery in the hospital 2

[0055] Cell culture medium: DMEM, Ham's F12 mixed medium, wherein, DMEM, Ham's F12 weight ratio 3: 1

[0056] Added factors: L glutamine (2mM), cholera toxin (0.1ug / ml), adenine (180uM), hydrocortisone (0.4ug / ml), insulin (10ug / ml), human epidermal growth factor (10ug / ml), transferrin (5ug / ml), triiodothyronine (2uM), CaCl 2 (1 mM).

[0057] (2) Cell isolation and culture steps

[0058] a) The skin samples were refrigerated and transported to the laboratory in DMEM culture medium, and the measurement area was 3cm 2 .

[0059] b) Wash the sample with PBS; wash the sample again after disinfection with iodine tincture and alcohol, cut off part of the dermis and chop it up

[0060] c) 0.05% trypsin cold digestion ...

example 3

[0065] Example 3 Foreskin skin after circumcision of healthy male, epidermal cells were expanded in vitro to form epidermis, and stained with hematoxylin and dopa.

[0066] (1) Materials and methods

[0067] Sample: 3cm of circumcised foreskin skin from healthy males discarded after surgery in the hospital 2 size

[0068] Skin medium: DMEM, Ham's F12 mixed medium, wherein, DMEM, Ham's F12 weight ratio 3:1

[0069] Added factors: L glutamine (2mM), cholera toxin (0.1ug / ml), adenine (180uM), hydrocortisone (0.4ug / ml), insulin (20ug / ml), human epidermal growth factor (10ug / ml), transferrin (5ug / ml), triiodothyronine (2uM), CaCl 2 (1 mM).

[0070] (2) Cell isolation and epidermal culture

[0071] a) Expansion of epidermal cells: the specific steps are described in Example 1

[0072] b) Separation and staining of the epidermal membrane: after 8 days, the cells were confluent, and after confluence, the skin was peeled off after continuing to culture for 4 days; after the epid...

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Abstract

The invention provides a serum-free culture system for epidermal cell culture. The serum-free culture system is composed of a basic culture medium, 1-5mM of amino acid, 0.01-0.5microgram / ml of cholera toxin, 0.1-100microgram / ml of somatomedin, 100-300NuM of adenine, 1-10microgram / ml of iron sources, 1-5NuM of triiodothyronine and calcium salts. The basic culture medium is free of serum and applicable to both trophoblast culture and trophoblast-free culture. Extracted primary cells are large in number and high in activity, epidermal cells are high in attachment rate, quick in multiplication and low in differentiation rate in a culture process, high cell activity is achieved after culture, and original cell functionality is kept.

Description

technical field [0001] The invention relates to a tissue engineering culture system, in particular to a serum-free culture system for epidermal cell culture. Background technique [0002] Epidermal cells are the cells located in the superficial layer of the skin and are mainly composed of epidermal forming cells and dendritic cells and other types of cells. Among them, epidermal progenitor cells exist in the basal layer of the epidermis and in the hair follicle. Epidermal cells play an important role in many cases. For example, in the construction of tissue engineered skin, epidermal progenitor cells can be introduced to cultivate the epidermal part of tissue engineered skin. In the pathological research and treatment of skin diseases, epidermal cells cultured in vitro can be used to It helps to study the pathogenesis of skin-related diseases and conduct wound repair and gene therapy through the response of epidermal cells to drugs. In addition, the healing effect of wounds...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
Inventor 刘洪李琳杨熙
Owner NOVAPRINT THERAPEUTICS SUZHOU CO LTD
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