High-efficiency preparation method of human skin tissue pluripotent stem cells
A pluripotent stem cell and skin technology, applied in the field of cell engineering, can solve the problems of low cell yield and low differentiation efficiency, and achieve the effect of strong stem cell activity, high differentiation efficiency, and maintaining stem cell properties.
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[0034] The invention discloses a highly efficient preparation method of human skin tissue pluripotent stem cells, the specific steps are:
[0035] 1. Coated bottle preparation:
[0036] (1) Dissolving human type IV collagen and diluting it to 0.1g / L with PBS to make a coating solution;
[0037] (2) Take 5ml of the coating solution and add it to the culture bottle, lay it flat, and put it in a refrigerator at 4°C overnight;
[0038] (3), discard the supernatant, and place it in an ultra-clean workbench to dry;
[0039] (4), the coated bottle is stored in a 4°C refrigerator for later use;
[0040] 2. Preparation of skin stem cells:
[0041] (5) Take cheek or chin skin tissue and remove fat tissue;
[0042] (6), cut the skin into small pieces of 0.5mm×0.5mm, rinse with PBS three times;
[0043] (7), the tissue block is placed in a sterile plate, with the dermis facing down, and digested overnight at 4°C with a digestion solution containing 1U / ml neutral protease II;
[0044...
Embodiment 1
[0059] The invention discloses a highly efficient preparation method of human skin tissue pluripotent stem cells, the specific steps are:
[0060] 1. Coated bottle preparation:
[0061] (1) Dissolving human type IV collagen and diluting it to 0.1g / L with PBS to make a coating solution;
[0062] (2) Take 5ml of the coating solution and add it to the culture bottle, lay it flat, and put it in a refrigerator at 4°C overnight;
[0063] (3), discard the supernatant, and place it in an ultra-clean workbench to dry;
[0064] (4), the coated bottle is stored in a 4°C refrigerator for later use;
[0065] 2. Preparation of skin stem cells:
[0066] (5), take cheek tissue, remove fat tissue;
[0067] (6), cut the skin into small pieces of 0.5mm×0.5mm, rinse with PBS three times;
[0068] (7), the tissue block is placed in a sterile plate, with the dermis facing down, and digested overnight at 4°C with a digestion solution containing 1U / ml neutral protease II;
[0069] (8), use opht...
Embodiment 2
[0084] The invention discloses a highly efficient preparation method of human skin tissue pluripotent stem cells, the specific steps are:
[0085] 1. Coated bottle preparation:
[0086] (1) Dissolving human type IV collagen and diluting it to 0.1g / L with PBS to make a coating solution;
[0087] (2) Take 5ml of the coating solution and add it to the culture bottle, lay it flat, and put it in a refrigerator at 4°C overnight;
[0088] (3), discard the supernatant, and place it in an ultra-clean workbench to dry;
[0089] (4), the coated bottle is stored in a 4°C refrigerator for later use;
[0090] 2. Preparation of skin stem cells:
[0091] (5), remove the skin tissue of the chin, and remove the fatty tissue;
[0092] (6), cut the skin into small pieces of 0.5mm×0.5mm, rinse with PBS three times;
[0093] (7), the tissue block is placed in a sterile plate, with the dermis facing down, and digested overnight at 4°C with a digestion solution containing 1U / ml neutral protease ...
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