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Method for culturing human limbal stem cell graft

A technology of limbal stem cells and limbal stem cells, which is applied in the field of culturing human limbal stem cell grafts using biological amniotic membrane as a carrier, can solve the problems of material collection, pollution in the processing process, troublesome preparation process, etc., and achieve the promotion of clinical application, convenient cultivation, High safety effect

Inactive Publication Date: 2012-07-18
上海交通大学附属第一人民医院
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Problems solved by technology

[0004] At present, most scholars use de-epithelialized amniotic membrane as a carrier to culture limbal epithelial cells 【2，3】 Wherein, the described epithelized amniotic membrane adopts the placental innermost layer fetus after cesarean section of healthy puerpera whose infectious index test is negative (including syphilis, HIV, hepatitis B, hepatitis C, hepatitis E and cytomegalovirus and bacteria detection). The membrane has been de-epithelized, but since the current detection method cannot detect the infected person in the incubation period, the possibility of long-term infection of the recipient cannot be completely avoided, and the preparation process is troublesome, and it may be contaminated during the sampling and processing process. It should be limited in clinical

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  • Method for culturing human limbal stem cell graft
  • Method for culturing human limbal stem cell graft
  • Method for culturing human limbal stem cell graft

Examples

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Embodiment 1

[0031] Tissue block adherence method: cut into 2.5×2.5cm 2 Biological amniotic membrane (epithelial side up) of the size of 100 mm, spread flat on the polycarbonate membrane of the insert culture dish, put the polycarbonate membrane into the culture dish of pre-inactivated 3T3 fibroblasts; under sterile conditions, the cornea Marginal tissue cut into 1mm 2 size, placed directly on the surface of the intact biological amniotic membrane; add 1ml of culture solution to the culture dish; add a few drops of culture solution to the surface of the tissue block to keep it moist; Fetal bovine serum, 5 μg / ml insulin, 20ng / ml rhEGF (recombinant human epithelial growth factor), 50IU / ml penicillin and streptomycin; place at 37°C, 5% CO 2 -Cultivate in an incubator with 95% air; replace the culture medium every 2 to 3 days; observe the growth of the cells under an inverted microscope every day;

[0032] The hematoxylin staining method of biological amniotic membrane and epithelial amnioti...

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Abstract

The invention belongs to the field of tissue engineering and relates to a method for culturing a human limbus stem cell graft by taking a biological amnion as a carrier. According to the method disclosed by the invention, the biological amnion is used as the carrier for culturing a limbus stem cell, wherein the biological amnion is a freeze-dried human amnion which is irradiated and sterilized by colbalt-60; the safety of the biological amnion is higher than that of denuded amniotic membrane; and the biological amnion is easy to preserve and convenient to acquire and is widely used clinically. The method disclosed by the invention has the advantages that an acceptor can be prevented from being infected for a long term; the preparation process is simple; the pollution in the material obtaining and treating process can be reduced; limbus epithelial cells can be cultured more safely and conveniently; and the clinical application of the human limbus stem cell graft is promoted.

Description

technical field [0001] The invention belongs to the field of tissue engineering and relates to a method for culturing human limbal stem cells, in particular to a method for cultivating human limbal stem cell grafts with biological amnion as a carrier. Background technique [0002] Tissue engineering is an emerging interdisciplinary subject in the field of cell transplantation. It applies the principles of engineering and life sciences to develop biological substitutes, and to improve cell functions lost due to disease through reconstruction through transplantation. The basic principle and method of tissue engineering to construct tissue engineered tissues and organs is to adsorb normal cells cultured and expanded in vitro on a biological scaffold material with good biocompatibility and which can be absorbed by the body to form The cell-biomaterial composite, and then, the cell-biomaterial composite is implanted into the body tissue; the diseased part of the organ forms a new...

Claims

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Application Information

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IPC IPC(8): C12N5/074
Inventor 刘焰黄洁
Owner 上海交通大学附属第一人民医院
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