Isolated culture method for enameloblast

A technology for the separation and cultivation of ameloblasts, applied in the field of separation and cultivation of ameloblasts, can solve the problems of cumbersome methods and high cost, and achieve the effects of improving purity, ensuring adherence and proliferation speed

Inactive Publication Date: 2013-08-21
SICHUAN UNIV
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  • Abstract
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Problems solved by technology

All of the above methods are cumbersome and costly

Method used

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  • Isolated culture method for enameloblast
  • Isolated culture method for enameloblast
  • Isolated culture method for enameloblast

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Embodiment 1

[0054] The tooth germ tissue was isolated from the fetal mouse jawbone tissue and cultured using the monoclonal method: the tooth germ was mechanically separated under a stereomicroscope under conventional aseptic conditions, after rinsing, shredding, and centrifugation, the tooth germ tissue was added with 20 times the volume of a mixture of 3mg / ml type Ⅰ collagenase and 3mg / ml dispase was digested at 37°C for 1.5h, during which it was properly shaken to fully digest, and the volume of medium equal to that of the mixture was added to neutralize and centrifuge to collect cells, and then the cells were collected with 0.05 % Trypsin was digested at 37°C for 5 minutes, and an equal volume of medium was added to the trypsin to neutralize, and the cells obtained after centrifugation were odontogenic cells.

[0055] The obtained odontogenic cells were inoculated on the vacuum-gas plasma-treated culture dishes with consistent surface chemical properties for adherent culture. Odontoge...

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Abstract

The invention relates to an isolated culture method for enameloblast, and particularly relates to a method for carrying out isolated culture on high-purity enameloblast through the combination of a method for separating dental germ tissues under a stereoscopic microscope and an enzyme digestion method. The method comprises the following steps: after mouse dental germs are separated under a stereoscopic microscope, carrying out primary culture on enameloblast by using a monoclonal method, carrying out purification by using culture solution type replacement and different digestion methods, observing cell morphology and growth conditions under an inverted microscope, and detecting the situations of cellular expressed amelogenin, amelogenin and cytokeratin 14 by using an immumofluorescence method. The growth of enameloblast subjected to primary culture is good, mesenchymal cells are significantly reduced after purification, and epithelial cells are subjected to lamellar growth so as to express keratin, amelogenin and amelogenin. The characteristics of in-vitro-cultured human epithelial enameloblast can be kept for a long time. The establishment of an in-vitro culture method for human dental epithelial cells provides a reliable seed cell source for the research on dental tissue engineering.

Description

technical field [0001] The invention relates to a method for separating and culturing ameloblasts, in particular to a method for separating and culturing high-purity ameloblasts by combining tooth germ tissue separation under a stereomicroscope with enzymatic digestion sorting. [0002] technical background [0003] Enamel, the most mineralized dental tissue, is composed of hydroxyapatite, approximately 96% minerals and the remaining 4% organic matter and water. The basic unit of enamel is the column. The weaving structure of the pillars strengthens the strength and the ability to resist fracture. However, this still does not prevent the enamel from being damaged by caries and trauma. Enamel is formed by ameloblasts of epithelial origin in the enamel organ of the developing tooth germ, and the enamel covers the dentin-pulp complex. Epithelial to ameloblast differentiation is regulated by interactions between epithelium and mesenchyme. The difference between enamel a...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 郑黎薇周学东万冕
Owner SICHUAN UNIV
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