Application of maduramicin ammonium in preparation of anti-breast cancer drugs
A kind of madomecin ammonium and anti-breast cancer technology, which is applied in the field of medicine, and can solve the problems such as no reports on the cytotoxic effect of madomecin ammonium on breast cancer
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Embodiment 1
[0035] According to the method in cell passage, six kinds of cells, MCF-10A, EpH4-Ev, MDA-MB-231, 4T1, CMT-U27 and canine primary breast cancer cells, were digested with trypsin respectively, and the above cells were divided into 5000 Cells / well were inoculated into 96-well cell culture plates, 100 μL per well, and cultured in a 37°C cell culture incubator. When the cells in each well grew to about 80%, the cells were treated with different concentrations (0μM, 0.0625μM, 0.0125μM, 0.25μM, 0.5μM, 1μM) of amdomicin ammonium for 24h, 48h, and 72h respectively, and 6 parallel replicates were designed for each group. hole, placed in CO 2 Continue culturing at 37°C in an incubator. After 24h, 48h, and 72h, 10 μL of CCK-8 solution was added to each well, and the incubation was continued for 1 hour in the incubator. The OD value at a wavelength of 450 nm was measured with a microplate reader.
[0036] The result is as figure 1 Shown: different concentrations of maduramicin ammoniu...
Embodiment 2
[0038] Select well-growing MDA-MB-231 cells and 4T1 cells, digest and centrifuge for passage, and when the cells grow to about 80% of the bottom of the culture dish, discard the original culture medium, and add different concentrations (0μM, 0.0625μM, 0.125μM, 0.25 μM, 0.5μM, 1μM) maduramicin ammonium sterile culture solution 10mL, continue to place in 5% CO 2routinely cultured in a cell culture incubator. Three time points of 24h, 48h, and 72h after administration were selected, and the cells were observed under a microscope and photographed.
[0039] The results show that: if figure 2 , 3 As shown, MDA-MB-231 cells and 4T1 cells were treated with different concentrations of maduramicin ammonium (0-1μM) for 24h, 48h and 72h respectively. With the increase of drug concentration and action time, the growth of breast cancer cells was inhibited. The activity gradually decreased, and at the same time, the cells gradually became round and shrunk, fell off from the culture dish ...
Embodiment 3
[0041] Digest well-grown MDA-MB-231 cells and 4T1 cells and inoculate 1×10 cells per well in a six-well plate 5 per cell to distribute the cells evenly across the plate. Place the culture plate in a 37°C cell culture incubator. On the next day, the original culture medium was discarded, and 2 mL of maduramicin ammonium sterile culture medium containing different concentrations (0 μM, 0.0625 μM, 0.25 μM, 1 μM) was added to each well, and the routine culture was continued in the cell culture incubator. After 48 hours of drug action on the cells, digest the cells with trypsin digestion solution (excluding EDTA), pipette into a cell suspension, transfer it to a 5mL centrifuge tube, place it in a centrifuge at 2000r / min for 5min, and then add pre-cooled PBS to wash the cell pellet Twice, collect 1-5×10 5 Add 500 μL of pre-cooled binding buffer to resuspend the cells, add 5 μL Annexin V-FITC and mix well, then add 5 μL propidium podide, mix well, and react for 15 minutes at room t...
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