Anti-ovarian cancer medicinal composition with synergistic effect and application thereof
A technology of synergy and composition, applied in the field of medical scientific research, to achieve the effect of good curative effect, good application prospect, and strong clinical application value
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Embodiment 1
[0042] Example 1 Cell Viability Determination
[0043] 1 Experimental method
[0044] Cell viability was determined using CCK-8 kit (Dojindo, Japan). Ovarian cancer OVCAR-3 cells were seeded into 96-well plates at a density of 6×10 per well 3cells. In time- and dose-dependent experiments, different concentrations of mangosteen (0, 5, 10, 20, 25, 30 and 50 μM) were added to equal individual wells. Empty (blank, no cells) and cells without drug treatment served as controls. Five wells were replicated for each concentration. After incubation for 24, 48 and 72 h, 10 μl of CCK-8 reagent was added to each well, and then the absorbance of optical density (OD) was read at 490 nm with a microplate reader (BioRad, Hercules, CA, USA) to evaluate the cells vitality.
[0045] For the combined drug experiment of mangosteen and cisplatin, we divided the cells into 3 groups: different doses (0, 5, 10, 20, 25 and 30 μM) of mangosteen were used alone, and different doses (0, 0.5, 1, 2, 4...
Embodiment 2
[0048] Example 2 Cell cycle analysis
[0049] 1 Experimental method
[0050] OVCAR-3 cells (concentration at 1×10 6 / well) were inoculated into 6-well plates and incubated with different concentrations of mangosteen (0, 10, 20 and 25 μM). After 48 hours of culture, the cells were collected by centrifugation, washed with cold phosphate-buffered saline (PBS), and then fixed with 70% cold ethanol overnight at 4°C. Subsequently, cells were incubated with 500 μl PBS containing 50 μg / ml propidium iodide (PI) and 0.1 mg / ml RNaseA (Sigma-Aldrich, USA) for 30 minutes at room temperature. Cellular distribution of the cell cycle was detected by FACSCalibur flow cytometry (BD Biosciences). Experiments were repeated at least three times.
[0051] 2 Experimental results
[0052] Mangosteen induces cycle arrest in ovarian cancer OVCAR-3 cells. OVCAR-3 cells were treated with different concentrations of mangosteen for 48 hours, and then cell cycle analysis was performed by flow cytometr...
Embodiment 3
[0056] Example 3 Detection of apoptotic cells by flow cytometry
[0057] 1 Experimental method
[0058] We used AnnexinV-FITC Apoptosis Detection Kit (BD Biosciences, USA) to detect apoptotic cells. Ovarian cancer OVCAR-3 cells in 2×10 6 Inoculated into a 6-well plate at a density of 1 / well, and after incubation for 24 hours, mangosteen and cisplatin were added to each well. The experiment was divided into three groups: mangostin alone group (20 or 25 μM), cisplatin alone group (2 or 4 μM) and combination group (20 μM mangostin plus 2 μM cisplatin). 3 wells were replicated at each concentration. After 48 hours of incubation with the drug, the cells were collected and washed once with PBS, and then 200 μl of binding buffer containing 5 μl of AnnexinV-FITC (fluorescein isothiocyanate) and 10 μl of PI (propidium iodide) staining solution was added . After incubation at room temperature for 15 minutes, apoptotic cells were detected by flow cytometry analysis. Experiments wer...
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