A kind of stem cell skin adhesive sheet and its preparation method and application
An adhesive sheet and stem cell technology, which is applied in the field of stem cell skin adhesive sheet and its preparation, can solve the problems of inability to repair skin function, fail to realize skin function repair, cumbersome and complicated preparation process, etc., so as to improve skin repair effect and reduce skin trauma quickly. , the effect of immune rejection is not easy
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[0032] In view of this, in a specific embodiment of the present invention, a kind of preparation method of stem cell skin adhesive sheet is provided, and described preparation method comprises:
[0033] S1. Separating and obtaining skin stem cells from skin tissue blocks;
[0034] S2, culture and passage expansion of skin stem cells;
[0035] S3. Lay the skin stem cells obtained through subculture and expansion on the cell membrane and incubate to obtain a cell paste sheet.
[0036] In yet another specific embodiment of the present invention, the specific method of step S1 is:
[0037] Take a piece of fresh skin tissue, soak it with 75% ethanol and 3% double antibody in PBS in turn; then chop the tissue, add digestive enzyme mixture to incubate and enzymolyze to obtain enzymatic hydrolysis solution I, and then add 0.25% trypsin to continue incubation Enzymolysis solution II was obtained, and then DNase I was added to continue incubation to obtain solution III; after the dige...
Embodiment 1
[0056] The specific steps of skin stem cell paste preparation:
[0057] 1. Isolation of skin stem cells: Take fresh skin tissue pieces weighing more than 0.1 g, treat them with 75% ethanol for 30 seconds, soak in PBS containing 3% double antibody for 3 minutes, and wash twice. Then mince the tissue with a scalpel blade, put it into a sterile EP tube, add 15ml of lyase mixture per gram of tissue (containing 4.0mg / L of lyase (dispase) and collagenase (collagenase), 37 ℃ water bath After incubation for 1 hour, mix well, then add 0.25% trypsin at a volume ratio of 1:3 and continue to incubate for 30 minutes, then add DNase I at a volume ratio of 1:1000 and continue to incubate for 5 minutes. Add an equal volume of stop solution (digestive enzyme inhibitor ) to terminate the digestion, gently pipette to obtain a single cell suspension, and filter the cell suspension with a 100 micron pore filter, take the filtered cells, resuspend and wash once with DMEM with serum, and then resusp...
Embodiment 2
[0060] Example 2 Transplantation test in vivo (nude mice) of skin sticking sheet
[0061] In order to verify that the prepared cell paste sheet has functional living skin stem cells, the prepared cell paste sheet was transplanted into nude mice with skin wounds to detect whether the wounds could heal and form complete skin.
[0062] (1) Take 8-week-old nude mice (nude / nude), after anesthesia ( figure 2 A), disinfect the skin on the back of the nude mouse, and use surgical scissors to cut off a circular area slightly smaller than the diaphragm to create a full-thickness skin wound;
[0063] (2) The prepared cell paste sheet was completely covered on the skin wound of nude mice, covered with Vaseline gauze, and bandaged (no suture needed) ( figure 2 B);
[0064] (3) The dressing tape can be removed after 10 days, and the wound has completely healed;
[0065] (4) After 1 month, raised human skin with pigment (higher than that of mouse skin) can be seen on the wound. And the...
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