Method of cultivating incisor cervical-loop epithelial cell of rodent by modified tissue culture technique
An improved tissue block method and epithelial cell technology, applied in the field of cell biology, can solve the problems of cumbersome steps, long time-consuming, unfavorable cell activity, etc., and achieve the effects of reducing cell damage, improving digestion efficiency, and shortening digestion time
Inactive Publication Date: 2012-07-25
FOURTH MILITARY MEDICAL UNIVERSITY
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Problems solved by technology
At present, the culture method of cervical ring epithelial cells is mainly enzymatic digestion method, which takes a long time and is not conducive to maintaining cell viability. Cells can be collected in stages to solve this problem, making the steps cumbersome
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[0026] The present invention will be described below in conjunction with specific examples, so that researchers in the field can better use this technology to culture cervical ring epithelial cells, but the examples given are not intended to limit the present invention. All experiments were performed in a clean bench.
[0027] 1) Primary culture of mouse incisor cervical ring epithelial cells
[0028] a. Choose a mouse 5-7 days after birth, soak it in 75% alcohol for 2-3 seconds, and kill the mouse by taking off the neck with a needle holder and tissue forceps, extract the mandible, and place it in sterile PBS.
[0029] b. After all the mandibles are extracted, Venus scissors and knotted tweezers separate the mandibular incisors. The rounded expansion of the root tip is the neck ring. Cut the labial neck ring tissue at the root tip and place it in 2ml with a concentration of 0.1%. Type I collagenase and 0.1% dispase II protease mixture, the digestion temperature is 37 degrees...
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The invention relates to the field of cell biology, particularly to a method of cultivating an incisor cervical-loop epithelial cell of a rodent by modified tissue culture technique. The method is characterized in that the method comprises the following steps: (1) collecting materials: extracting a cervical-loop tissue at the lip side of a mandible incisor root tip in the rodent; (2) digesting: digesting a tissue block in an enzyme mixed liquor, wherein the enzyme mixed liquor is an enzyme digestive liquid prepared from 0.1% dispase II (a protease) and 0.1% type I collagenase, the digestion temperature is 37 DEG C, and the digestion time is 15-20min; (3) inoculating: collecting the tissue block after centrifugation of the digestive liquid, and inoculating onto a culture plate; and (4) purifying: removing fibroblasts to obtain purified cervical-loop epithelial cells. According to the method, during the primary cell culture, the enzymatic digestion temperature is 37 DEG C, the digestion time is shortened, the time is saved, the method facilitates cell growth out of the tissue blocks, and the cells are not damaged.
Description
technical field [0001] The invention relates to the field of cell biology, in particular to a method for cultivating rodent incisor cervical epithelial cells by an improved tissue block method. Background technique [0002] Dental trauma, dental caries, periodontal disease, and genetic diseases can all lead to tooth loss, directly affect the masticatory function and aesthetics of patients, and have a negative impact on patients' psychology. The current methods of tooth restoration mainly rely on the remaining teeth and periodontal tissue, causing damage to healthy teeth and periodontal tissues to varying degrees. Regenerated teeth have good biocompatibility and do not damage adjacent teeth and periodontal tissue, so tooth regeneration has become a research hotspot in recent years. Rodent incisors can grow continuously throughout life due to the formation of stem cell niches in the cervical ring at the root tip, and are often used as animal models for tooth regeneration stu...
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IPC IPC(8): C12N5/071
Inventor 邢向辉王小竞张陶涛
Owner FOURTH MILITARY MEDICAL UNIVERSITY