Medicine box for wound healing and preparation method thereof
A technology of wound repair and medicine box, which is applied in the field of biomedicine, can solve the problems of high price of acellular dermal matrix, difficulty in popularization and application in primary hospitals, and long overall treatment time, so as to accelerate wound healing rate, simple and quick preparation method, and improve Healing Mass Effect
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Embodiment 1
[0048] Embodiment 1, the preparation of the kit for wound repair
[0049] (1) Rapid separation and extraction of epidermal basal cell suspension:
[0050] A. Digestion: Take a 1cm×2cm blade-thick skin piece with a thickness of about 0.2mm and place it in a petri dish, immediately add TrypLESelect (10×) digestion solution and digest at 37°C for 15 minutes; among them, TrypLE Select (10×) The solution was purchased from Thermo Fisher Scientific (article number: A1217701), and was preheated to 37°C before use;
[0051] B. Separation: Add lactated Ringer's solution to rinse the skin and stop digestion, separate the epidermis and dermis with tweezers, and gently scrape the EBCs from the basal layer of the epidermis with a sterile scalpel, draw the EBCs with a 1ml syringe and a round needle, and collect EBCs were then diluted with 2ml lactated Ringer's solution to obtain a cell suspension;
[0052] C. Extraction and filtration: Filter the above cell suspension into a 50ml centrifu...
Embodiment 2
[0054] Embodiment 2, the using method of the medicine box that the present invention is used for wound repair:
[0055] S1. Spray fibronectin antibacterial solution on the fresh granulation wound bed and the wound bed in the skin extraction area after the patient has completely removed the scar until it evenly covers the wound bed (each ml of fibronectin antibacterial solution covers about 10 cm) 2 Wound), let stand for 30min;
[0056] S2. Spray the EBCs suspension on the wound bed covered with fibronectin, and the cell suspension can be evenly attached to the wound bed without obvious loss.
[0057] EBCs can be prepared by separating and extracting skin slices, or they can be artificially cultured cells. When the cells are artificially cultured, the cells need to be routinely cultured in advance, and then the cells are taken out, centrifuged to remove the supernatant, and then resuspended with lactated Ringer's solution to obtain a cell suspension, which is divided into fibr...
experiment example 1
[0058] Experimental example 1. Identification of ESCs purity
[0059] 1. Rapid separation and extraction of EBCs: The method is the same as that in Example 1. After the EBCs cell suspension is filtered, centrifuged, and supernatant discarded, it is resuspended with K-SFM medium.
[0060] 2. Rapid adsorption and separation of ESCs:
[0061] (1) Coating: ESCs were separated and extracted from EBCs by the differential attachment method, and the cell culture dish was coated with 20 μg / ml FN at 37°C for 30 minutes in advance;
[0062] (2) Seed plate: the separated EBCs were separated by 10 5 / cm 2 Seed the plate and culture it statically at 37°C for 10-20 minutes;
[0063](3) The non-adhered cells were washed away with medium, and the adhered cells were ESCs.
[0064] 3. Identification of ESCs purity by immunofluorescence
[0065] (1) Cell climbing sheet: put 10 4 The EBCs obtained by rapid separation and extraction were inoculated onto 20 μg / ml FN-coated 4-chamber immunofluo...
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