Stem cell exosome microneedle patch for removing freckles and wrinkles and preparation method of stem cell exosome microneedle patch
A stem cell and exosome technology, used in microneedles, medical preparations with inactive ingredients, and medical preparations containing active ingredients, etc., can solve the problem of poor individual targeting, serious product homogeneity, and single active ingredients. It can reduce the deposition of scars and pigments, reduce melanin, and inhibit the activity of tyrosinase.
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Embodiment 1
[0048] Example 1 Preparation of stem cell exosomes
[0049] 1. Isolation and culture of umbilical cord mesenchymal stem cells
[0050] Take 15-20cm long neonatal umbilical cord, require obstetrics and gynecology healthy full-term caesarean section fetus, pregnant women HBV antigen, anti-HCV antibody, anti-HIV antibody, anti-Treponema pallidum antibody, mycoplasma, anti-cytomegalovirus antibody and other tests are all negative. Treat within 6 hours, wash with PBS under sterile conditions until there is no blood, cut it longitudinally, remove the blood vessels and cut it into pieces of tissue with a size of 1mm×1mm×1mm, add 0.5ml of umbilical cord tissue suspension to a 75cm2 culture bottle, and add 10ml at the same time Serum-free complete medium, cultured in a 37°C, 5% CO2 incubator. Change the medium in half on the 3rd day and the full volume on the 7th day. Observe under an inverted microscope. When the cells grow to 80% confluent, digest with 0.05% trypsin and passage at a...
Embodiment 2
[0055] Example 2 Identification of stem cells and detection of exosomes
[0056] 1. Identification of umbilical cord mesenchymal stem cells
[0057] (1) Take 1 bottle of T175 bottle of umbilical cord mesenchymal stem cells with a confluence of 80% to 90% (cell number 1×10 7 ), digest with 5ml 0.05% trypsin for 3-5min; when most of the cells shrink and become round under the microscope, add 2mL serum-containing medium to stop the digestion, blow and beat the cells and transfer them to a 15mL centrifuge tube for centrifugation at 800rpm, centrifuge The time is 5 minutes;
[0058] (2) Cell preparation: Discard the supernatant, add 2mL PBS to suspend the pellet, and count the cells; 5 Distribute the cell suspension evenly into 1.5mL Ep tubes, centrifuge at 800rpm for 5min; discard the supernatant, add 100μL of PBS to the pellet to suspend;
[0059] (3) Antibody incubation: Add 1 μL of the corresponding flow-type antibody to each tube, incubate in the dark for 40 minutes, wash w...
Embodiment 3
[0064] Example 3 Preparation of Stem Cell Exosome Microneedle Patch
[0065] 1. Preparation of needle tip solution (exosome repair solution)
[0066] (1) Composition:
[0067] Exosomes (2.5×10 14 / L);
[0068] Astaxanthin (15g / L);
[0069] Trehalose (40g / L);
[0070] Chitosan (7.5g / L);
[0071] Bovine serum albumin (2g / L);
[0072] Phosphate buffer (0.015mol / L) was used to adjust the pH value to 7.2.
[0073] (2) The solution prepared in the above steps was mixed according to the formula to prepare an exosome repair solution, and then filtered through a 500-mesh cell sieve to obtain a microneedle tip solution.
[0074] 2. Preparation of needle solution
[0075] The needle solution was prepared by PVP K30 (2000g / L), PEG400 (20g / L), squalane (20g / L) and sterile water 500μl.
[0076] 3. Preparation of waterproof backing layer
[0077] (1) Backing layer formula: 4-8 parts of film material polyvinyl alcohol, 2-4 parts of thickener sodium carboxymethyl cellulose, 2-8 parts...
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