Protection effect of cyanidin-3-glucoside for UVB (ultraviolet B)-induced HaCaT cell injury
A technology of glucoside and cyanidin, which is applied in the direction of medical preparations containing active ingredients, cosmetic preparations, skin care preparations, etc., to protect DNA from damage, improve survival rate, prevent and treat skin light damage effect
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Embodiment 1
[0051] Example 1 Study of cyanidin-3-glucoside interfering with ROS / COX-2 pathway in HaCaT cells damaged by UVB
[0052] 1. Experimental method
[0053] 1. HaCaT cell culture
[0055] 1) Take out the cryopreservation tube containing HaCaT cells from the -80°C refrigerator, put it into warm water at 37°C immediately, and shake it from time to time to make it evenly heated and melt as soon as possible;
[0056] 2) When the cryopreservation solution is completely dissolved, take out the cryopreservation tube, suck out the cell suspension in a sterile table, inject it into 9ml of cell culture medium, blow it evenly, and centrifuge at 1000rpm×5min;
[0057] 3) Discard the supernatant, add 6ml medium, pipette the discrete cells, transfer to 10mm Petri dish, place in 5% CO 2 , Cultured in a 37°C incubator.
[0058] (2) Cell culture
[0059] 1) Cells were grown in DMEM medium containing 10% FBS and placed in 5% CO 2 , Cultivate in a 37°C incubator; c...
Embodiment 2C3
[0172] The effect of embodiment 2C3G on the apoptosis of HaCaT cells after UVB irradiation
[0173] 1. Apoptosis detection——AnnexinV-FITC / PI double staining flow cytometry detection
[0174] The principle of AnnexinV-FITC / PI double staining method for detecting cell apoptosis: in the early stage of cell apoptosis, phosphatidylserine (phosphalidylserine, PS) located on the inner side of the cell membrane can migrate to the outer side of the cell membrane, and AnnexinV, as a phospholipid-binding protein, can inhibit PS It has a high binding force, so the fluorescence intensity of AnnexinV-labeled fluorescein (FITC) can reflect early apoptotic cells; at the same time, because the PS of necrotic cells is also exposed outside the cell membrane, and is highly stained to PI, the combined use of PI to reject Apoptotic (early apoptosis) and necrotic (late apoptosis) cells can be distinguished by staining. On the scatter diagram of flow cytometry analysis, AnnexinV and PI double nega...
Embodiment 3C3
[0180] Effect of embodiment 3C3G on the mitochondrial membrane potential of HaCaT cells after UVB irradiation
[0181] 1. Mitochondrial membrane potential (Δψm) detection—JC-1 staining flow cytometry detection
[0182] The principle of JC-1 detecting the change of mitochondrial membrane potential: JC-1 is a cationic lipid fluorescent dye, which has two states of monomer and polymer. exist in multimeric form. In normal cells, due to the polarity of the mitochondrial membrane potential (Δψm), JC-1 is rapidly absorbed into the mitochondria depending on the polarity of Δψm, and forms multimers in the mitochondria due to its increased concentration, and the multimers emit The light is red fluorescence, which can be detected by the red (FL-2) channel of the flow cytometer; when cells undergo apoptosis, the mitochondrial membrane potential is depolarized, resulting in the release of JC-1 from the mitochondria, reducing its concentration , so it exists in the form of a monomer and...
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