Preparation method of blue-dyed amniotic basement membrane coated with fibrinogen
A technology of fibrinogen and basement membrane, which is applied in the field of preparation of blue-stained amniotic membrane basement membrane
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Embodiment 1
[0044] A kind of preparation method of the blue-dyed amniotic membrane basement membrane of coating fibrinogen, comprises the following steps:
[0045] S1, preparation of amnion basement membrane: the amnion is washed with sterile phosphate-buffered saline PBS containing 0.3wt% ofloxacin to remove blood and impurities,
[0046] The amnion was mixed with trypsin at a mass concentration of 2.5%, incubated at 37°C for 6 hours, and the serosa and part of the matrix of the amnion were scraped off with a cell scraper.
[0047] Then soak the amniotic membrane in gelatinase IV with a mass concentration of 0.3 mg / ml, incubate at 37°C for 3 hours, digest the remaining amniotic matrix and dense layer, and wash twice with sterile PBS,
[0048] Then soak the amniotic membrane in 0.02% ethylenediaminetetraacetic acid EDTA, incubate at 37°C for 2 hours, then scrape gently to remove the amniotic membrane epithelium, obtain the treated amniotic membrane basement membrane, and place it on Dulb...
Embodiment 2
[0053] This example is basically the same as Example 1, the difference is that the specific steps of trypan blue staining of amniotic membrane basement membrane are as follows:
[0054] S2-1-1: Soak the amniotic basement membrane in trypan blue staining agent for 3 minutes;
[0055] S2-1-2: Take out the soaked amniotic basement membrane and soak in the perfusion solution for 3 minutes;
[0056] S2-1-3: drying the stained amniotic basement membrane under sterile conditions at 32°C.
[0057] Among them, the viscoelastic agent is a colorless transparent gel-like solution composed of high-purity sodium hyaluronate and physiological buffer balance salts. % by weight, 0.05% by weight of potassium chloride, 0.05% by weight of calcium chloride dihydrate, 0.03% by weight of magnesium chloride hexahydrate, 0.3% by weight of sodium acetate trihydrate, 0.17% by weight of sodium citrate dihydrate, and the balance is water.
Embodiment 3
[0059] This embodiment is basically the same as Embodiment 2, the difference being that the dyeing parameters are different:
[0060] S2-1-1: Soak the amniotic basement membrane in trypan blue staining agent for 4 minutes;
[0061] S2-1-2: Take out the soaked amniotic basement membrane and soak in the perfusion solution for 4 minutes;
[0062] S2-1-3: drying the stained amniotic basement membrane under sterile conditions at 33°C.
[0063] Among them, the viscoelastic agent is a colorless transparent gel-like solution composed of high-purity sodium hyaluronate and physiological buffer balance salt, and the composition content and ratio of the perfusate are: mannitol 2.5wt%, glucose 1.4wt%, 0.8wt% of sodium, 0.06wt% of potassium chloride, 0.05wt% of calcium chloride dihydrate, 0.03wt% of magnesium chloride hexahydrate, 0.3wt% of sodium acetate trihydrate, 0.17wt% of sodium citrate dihydrate, and the balance is water.
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