Method for preparing bovine cornea stroma from fresh bovine cornea and application method

A technology of bovine cornea and stroma, applied in medical science, tissue regeneration, prosthesis, etc., can solve the problems of scaffold structure damage, increase in the number of patients, and inability to apply clinically, and achieve low cost, short operation time, and no antigenicity Effect

Inactive Publication Date: 2016-06-08
北京智慧细胞生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The 2006 "Second National Sampling Survey of Disabled Persons" showed that there were about 4 million blind patients due to corneal diseases, and more than 100,000 new patients were added every year, of which about 2 million could recover their eyesight through corneal transplantation, but every year The number of corneal transplants in China is 4,000-5,000. A large number of patients are blind due to lack of donor sources. About 3 million patients are "waiting for the rice to cook." The provision of not allowing the use of the remains of executed prisoners has greatly reduced the supply of corneas. The number of patients waiting in line for corneal donors will increase significantly each year
[0003] In terms of corneal carrier scaffolds, although domestic and foreign scholars have conducted a large number of experimental studies on natural biomaterials and composite m

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  • Method for preparing bovine cornea stroma from fresh bovine cornea and application method
  • Method for preparing bovine cornea stroma from fresh bovine cornea and application method
  • Method for preparing bovine cornea stroma from fresh bovine cornea and application method

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Embodiment 1

[0033] The present invention provides a kind of method utilizing fresh bovine cornea to prepare bovine corneal stroma, comprising the following steps:

[0034] S1. Take the fresh bull's eyeball taken within no more than 8 hours, rinse it with normal saline, cut out a 200-micron corneal slice with a microkeratine trephine, and put it into a 0.6% NaCl hypotonic solution for 30 minutes.

[0035] S20. Use a scalpel to scrape off the corneal epithelial layer, retaining the Bowman's membrane, and operate gently until the Bowman's membrane layer is scraped. With the help of surgical scissors, tear off the Descemet's membrane and the endothelial layer to obtain the bovine corneal stroma including the Descemet's membrane and part of the stroma. Place the bovine corneal stroma slices in a 50ml cryopreservation tube filled with PBS, freeze in a -80°C refrigerator for 1 hour, then thaw at 37°C for 15 minutes, repeat freezing and thawing three times to break the cell wall, so that the bovi...

Embodiment 2

[0043] The present invention provides a kind of method utilizing fresh bovine cornea to prepare bovine corneal stroma, comprising the following steps:

[0044] S1. Take the fresh bull's eyeball taken within no more than 12 hours, wash it with normal saline, cut out a 300-micron corneal slice with a microkeratine trephine, and put it into a 0.7% NaCl hypotonic solution for 45 minutes.

[0045] S20. Use a scalpel to scrape off the corneal epithelial layer, retaining the Bowman's membrane, and operate gently until the Bowman's membrane layer is scraped. With the help of surgical scissors, tear off the Descemet's membrane and the endothelial layer to obtain the bovine corneal stroma including the Descemet's membrane and part of the stroma. Place the bovine corneal stroma slices in a 50ml cryopreservation tube filled with PBS, freeze in a -80°C refrigerator for 2 hours, then thaw at 37°C for 45 minutes, and repeat freezing and thawing 4 times to break the cell wall to make the bovi...

Embodiment 3

[0053] The present invention provides a kind of method utilizing fresh bovine cornea to prepare bovine corneal stroma, comprising the following steps:

[0054] S1. Take the fresh bull's eyeball taken within no more than 12 hours, wash it with normal saline, cut out a 400-micron corneal slice with a microkeratine trephine, and put it into a 0.8% NaCl hypotonic solution for 60 minutes.

[0055] S20. Use a scalpel to scrape off the corneal epithelial layer, retaining the Bowman's membrane, and operate gently until the Bowman's membrane layer is scraped. With the help of surgical scissors, tear off the Bowman's membrane and endothelial layer to obtain the bovine corneal stroma including the Bowman's membrane and part of the stroma layer, place the bovine corneal stroma slices in a 50ml cryovial containing PBS, and store at -80°C Freeze in the refrigerator for 3 hours, then thaw at 37°C for 60 minutes, repeat freezing and thawing three times to break the cell wall, so as to break t...

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Abstract

The invention provides a method for preparing a bovine cornea stroma from a fresh bovine cornea. The method comprises the steps that a cornea piece is cut from a fresh bovine eyeball and then steeped in a hypotonic solution; the bovine cornea stroma containing a lamina elastica anterior and part of a stroma layer is obtained from the cornea piece, and is frozen and thawed repeatedly to break cells; the bovine cornea stroma is bleached with a buffer solution, steeped in a cell removing reagent and then washed with ultrapure water in sequence; the bovine cornea stroma is put into another cell removing reagent for steeping, and then washed with ultrapure water again; the bovine cornea stroma is put into a dehydrating agent to be steeped and then irradiated with cobalt-60, and then the prepared bovine cornea stroma is dried and stored. The physical method and low-toxicity regents are adopted in the method, and time is very short, so that the obtained bovine cornea stroma is completely preserved, has the advantages of being good in transparence and compact in structure, and is close to the natural feature of the human cornea.

Description

technical field [0001] The invention relates to a method for preparing bovine corneal stroma, in particular to a method for preparing bovine corneal stroma by utilizing fresh bovine cornea and an application method. Background technique [0002] Corneal disease is the fourth leading cause of blindness in the world. According to the World Health Organization, corneal disease is the fourth leading cause of blindness after cataracts, glaucoma, and age-related macular degeneration, accounting for about 5.1%. The 2006 "Second National Sampling Survey of Disabled People Statistical Results" showed that there were about 4 million blind patients due to corneal diseases, and more than 100,000 new patients were added every year, of which about 2 million could recover their eyesight through corneal transplantation. The number of corneal transplants in China is 4,000-5,000, and a large number of patients are blind due to lack of donor sources. About 3 million patients are "waiting for t...

Claims

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Application Information

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IPC IPC(8): A61L27/36A61L27/50
CPCA61L27/3604A61L27/3683A61L27/3687A61L27/50A61L2430/16A61L2430/40
Inventor 韩斌张诚张旭
Owner 北京智慧细胞生物科技有限公司
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