FSHR (follicle-stimulating hormone receptor)-targeted tumor treatment polypeptide and application thereof
A tumor treatment, targeting technology, applied in the field of biomedicine
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Embodiment 1
[0057] Detecting the expression of FSHR in several kinds of cells by Western Blot
[0058] Human prostate cancer cell line PC3, human cervical cancer cell line Hela, human ovarian cancer cell line SKOV3, human breast cancer cell line MCF7, human embryonic kidney cell 293, and mouse fibroblast L929 were purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences.
[0059] Male BALB / c-nu nude mice, weighing 18-19 g, were purchased from Jiangsu Changzhou Cavens Experimental Animal Co., Ltd., SPF grade, and aged 5 weeks. Certificate number: NO.0026328, license number: SCXK (Su) 2011-0003.
[0060] The cell culture methods of PC3, MCF7, Hela, PC3, 293, SKOV3, and L929 used in the experiment were the same, at 37°C, 5% CO 2 Cultured and subcultured in a constant temperature incubator, cultured with DMEM medium containing 10% fetal bovine serum, 100U / ml penicillin, and 100U / ml streptomycin.
[0061] Collect cells, use RIPA reagent containing protease inhibitors, extract...
Embodiment 2
[0065] Determination of peptide FSH by MTT method 33-53 -The effect of IIKK on the viability of various cells
[0066] Cells grown in logarithmic phase were divided into 5×10 3 The density of cells / well was seeded in a 96-well plate. After culturing overnight, after the cells adhered to the wall, each well was added with different concentrations of FSH 33-53 - 100 μl of culture medium of IIKK, IIKK and cisplatin (concentration gradient is 0, 1, 2, 5, 10, 15, 20, 40, 50 μM), and three replicate wells are set for each concentration. After incubation for 24 hours, add 40 μl of MTT solution with a concentration of 2 mg / ml to each well and continue culturing, and discard the supernatant after 4 hours; add 150 μl of DMSO to each well and shake for 10 minutes to fully dissolve the crystals. A microplate reader was used to measure the OD value of the sample at a wavelength of 570 nm. Calculate the survival rate, draw the survival curve, and find the IC 50 . Survival rate=(OD val...
Embodiment 3
[0074] FSH 33-53 - Effect of IIKK on PC3 cell cycle
[0075] The cell cycle refers to the activity process of cells from the end of the previous division to the end of the next division, which is divided into two stages: interphase and division phase. Among them, the interphase is divided into G1 phase (pre-DNA synthesis), S phase (DNA synthesis phase), and G2 phase (late DNA synthesis phase). M phase is the mitotic phase. The G0 phase is the cell dormancy phase. Propidium iodide (PI) is a nuclear staining reagent that can stain DNA and RNA, and can release red fluorescence after intercalating DNA or RNA. Although PI cannot pass through the living cell membrane, it can pass through the damaged cell membrane and stain the nucleus. Cold ethanol can increase cell membrane permeability. RNA is cleaved with RNase, allowing PI to enter the cell and selectively bind to DNA. The fluorescence of the dye in the cell is proportional to the DNA content in the nucleus. The relative D...
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