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Amniotic membrane long-term preserving fluid and preparation method thereof

A long-term preservation and preservation solution technology, applied in the field of amniotic membrane long-term preservation solution and its preparation, can solve the problems of not achieving amniotic cell activity, depleting amniotic cell energy, affecting amniotic cell survival, etc., maintaining amniotic cell activity and optimizing amniotic membrane preservation. conditions, the effect of reducing the spread of the virus

Active Publication Date: 2011-07-27
周海华
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it has the following shortcomings: serum components are added to the preservation solution, which increases the chance of virus transmission between the amniotic membrane and the preservation solution; the ingredients are mainly a variety of viscous agents, and only 0.9-1.2% of ordinary DMEM is added As a component of the preservation solution, the culture medium is far from meeting the requirements for maintaining the activity of amnion cells; without adding cell nutrients, the cells will soon undergo apoptosis or even necrosis, but the storage time of amnion is only slightly prolonged; without adding vascular expansion agent, it is easy to make amnion cells and tissue swelling, thereby causing amnion cell apoptosis and collagen destruction; although the added dexamethasone can promote cell division and differentiation, it consumes the energy of some amnion cells, affects the survival of amnion cells, and may also be immersed in the amnion lining that affects the function of the amniotic membrane

Method used

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  • Amniotic membrane long-term preserving fluid and preparation method thereof
  • Amniotic membrane long-term preserving fluid and preparation method thereof
  • Amniotic membrane long-term preserving fluid and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Based on 1000ml preservation solution, including 20g of hydroxyethyl starch, 25g of chondroitin sulfate and 10g of low molecular weight dextran, 10ml of 100mM non-essential amino acid, 10ml of 100mM sodium pyruvate, 10ml of 500mM ascorbic acid, 25ml of 1M HEPES, 10ml of 10% tobramycin , DMEM / F12 culture medium is 935ml. The pH value is 7.2-7.4, and the osmotic pressure is 350-380mOsm / L.

[0045] Preparation method 1: Add 25g of chondroitin sulfate, 10g of low-molecular dextran, and 20g of hydroxyethyl starch to 10ml of 100mM non-essential amino acids, 10ml of 100mM sodium pyruvate, 10ml of 500mM ascorbic acid, 25ml of 1M HEPES, and 10ml of 10% tobramycin and 935ml DMEM / F12 culture medium, mix evenly, adjust the pH value to 7.2-7.4, and the osmotic pressure to 350-380mOsm / L, filter through a 0.2μm membrane to sterilize in 100ml aliquots, and store at 4°C to obtain this preservation solution.

[0046] Preparation method 2: Mix 10ml of 100mM non-essential amino acid, 10ml...

Embodiment 2

[0048] Based on 1000ml preservation solution, including 20g of hydroxyethyl starch, 25g of chondroitin sulfate and 10g of low molecular weight dextran, 10ml of 100mM non-essential amino acid, 10ml of 100mM sodium pyruvate, 10ml of 500mM ascorbic acid, 25ml of 1M HEPES, 10ml of 10% tobramycin , KSFM culture medium is 935ml. The pH value is 7.2-7.4, and the osmotic pressure is 350-380mOsm / L.

[0049] Preparation method 1: Add 25g of chondroitin sulfate, 10g of low-molecular dextran, and 20g of hydroxyethyl starch to 10ml of 100mM non-essential amino acids, 10ml of 100mM sodium pyruvate, 10ml of 500mM ascorbic acid, 25ml of 1M HEPES, and 10ml of 10% tobramycin and 935ml KSFM culture medium, mix evenly, adjust pH to 7.2-7.4, osmotic pressure to 350-380mOsm / L, filter through 0.2μm membrane to 100ml aliquots and store at 4°C to obtain this preservation solution.

[0050] Preparation method 2: Mix 10ml of 100mM non-essential amino acid, 10ml of 100mM sodium pyruvate, 25ml of 1M HEPE...

Embodiment 3

[0052] Based on 1000ml preservation solution, including 20g of hydroxyethyl starch, 25g of chondroitin sulfate and 10g of low molecular weight dextran, 10ml of 100mM non-essential amino acid, 10ml of 100mM sodium pyruvate, 10ml of 500mM ascorbic acid, 25ml of 1M HEPES, 10ml of 10% tobramycin , 1640 culture medium is 935ml. The pH value is 7.2-7.4, and the osmotic pressure is 350-380mOsm / L.

[0053] Preparation method 1: Add 25g of chondroitin sulfate, 10g of low-molecular dextran, and 20g of hydroxyethyl starch to 10ml of 100mM non-essential amino acids, 10ml of 100mM sodium pyruvate, 10ml of 500mM ascorbic acid, 25ml of 1M HEPES, and 10ml of 10% tobramycin and 935ml of 1640 culture medium, mix evenly, adjust the pH value to 7.2-7.4, and the osmotic pressure to 350-380mOsm / L, filter and sterilize through a 0.2μm membrane, and store in 100ml aliquots at 4°C to obtain this preservation solution.

[0054] Preparation method 2: Mix 10ml of 100mM non-essential amino acid, 10ml of ...

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Abstract

The invention relates to an amniotic membrane long-term preserving fluid which is characterized in that 1000ml of preserving fluid contains 10g-200g of chondroitin sulfate and low-molecular-weight dextranum, 10g-50g of hydroxyethyl starch, 5ml-20ml of amino acid, 5ml-20ml of tobramycin, 10ml-50ml of HEPES, 5ml-30ml of ascorbic acid, 5ml-20ml of sodium pyruvate, and 860ml-970ml of culture solution. A preparation method of the amniotic membrane long-term preserving fluid comprises the following steps: adding the chondroitin sulfate, the low-molecular-weight dextranum, the amino acid, the tobramycin, the hydroxyethyl starch, the ascorbic acid and the sodium pyruvate into the culture solution and then uniformly mixing the mixture; using 1%-5% of HEPES to adjust pH value till the pH value is 7.2-7.4; using an osmotic pressure buffering agent to adjust the osmotic pressure till the osmotic pressure is 350-380mOsm / L; and performing membrane filtration and degerming, thereby acquiring the amniotic membrane long-term preserving fluid.

Description

technical field [0001] The invention relates to an amniotic membrane preservation solution, in particular to a long-term amniotic membrane preservation solution and a preparation method thereof. Background technique [0002] Amniotic membrane (thickness about 0.02 ~ 0.05mm) is derived from the embryo, is a transparent, no nerve, blood vessels and lymphatic vessels, low antigenicity and tough tissue, composed of epithelial cell layer, basement membrane, stroma layer, fibroblast layer , Sponge layer composition. Amniotic membrane plays an important role in many fields of ophthalmology, such as promoting epithelial repair, healing and differentiation, inhibiting inflammation, anti-angiogenesis, inhibiting fibrosis and scarring, and preventing synechia. With the increasing amount of amniotic membrane, the preservation of amniotic membrane, especially the preservation of the active ingredients of amniotic membrane, has become a hot spot in the field of ophthalmology. [0003] T...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
Inventor 周海华
Owner 周海华
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