Tumor vaccine, and preparation method and application thereof
A tumor vaccine and tumor technology, applied in the field of medicine, can solve the problems such as no lactic acid, no tumor vaccine, etc., and achieve the effects of prolonging the survival time, improving the survival rate, and inhibiting growth.
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Embodiment 1
[0056] Effect of embodiment 1 lactic acid on tumor cell growth
[0057] 1. Experimental method
[0058] The effect of lactic acid on tumor cell proliferation was analyzed by MTT assay. Collect the E.G7 cells grown in logarithmic phase, centrifuge to discard the supernatant, resuspend and adjust the cell concentration to 2×10 4 / ml. Divide the resuspended E.G7 cells into 5 groups, add 0mmol / L, 5mmol / L, 10mmol / L, 20mmol / L, 30mmol / L lactic acid respectively. The above-prepared cell suspension was put into a sterile 96-well plate with a volume of 200 μl per well, and each group had 3 replicate wells, and sterile physiological saline was added to the edge wells to fill. The above-mentioned 96-well plate was placed in a 5% CO2 constant temperature cell culture incubator at 37° C. for 24 hours. Then, 20 μl of MTT solution (5 mg / ml) was added to each well, and placed in an incubator for incubation for 4 hours. Take out the 96-well plate, centrifuge at 250g for 5min, remove the su...
Embodiment 2
[0062] Example 2 The phagocytosis of dendritic cells on the tumor whole cell vaccine of the present invention
[0063] 1. Experimental method
[0064] In order to study whether the phagocytosis of DCs to the whole tumor cell vaccine of the present invention is enhanced, in this experiment, the tumor cells stimulated by different concentrations of lactic acid and irradiated were stained with PHK67, then mixed with DCs and cultured overnight, and DCs were screened out by flow cytometry. After that, the percentage of CD11c and PHK67 double positive cells was detected. The specific experimental method is as follows:
[0065] E.G7 cells were stimulated according to the concentration gradient of lactic acid, including: 0mmol / L, 5mmol / L, 10mmol / L, 20mmol / L, and 30mmol / L, and irradiated. Diluent C (100μlper 2×10 5 cells) to resuspend the cells, add 0.4 μl of PKH67 staining solution to every 100 μl of Diluent C, pipette repeatedly 10 times, and incubate at room temperature for 2-5 m...
Embodiment 3
[0070] Example 3 Prevention experiment and treatment experiment of tumor whole cell vaccine of the present invention
[0071] 1. Experimental method
[0072] 6-8 weeks old, 16-18 g female C57BL / 6J mice (for E.G7 mouse T-cell lymphoma model) or BALB / c mice (for 4T1 mouse breast cancer model) were selected for this experiment. Each model was divided into 3 groups with 10 mice in each group. The specific groups are: 1) normal saline group (saline): 200 μl of normal saline; 2) control group: 1×10 6 Cells / 100μl E.G7 cells irradiated with 75Gy 200μl or 1×10 6 Cells / 100μl 200μl of 4T1 cells irradiated with 50Gy; 3) Experimental group: 1×10 6 Cells / 100 μl E.G7 cells 200 μl or 1×10 cells stimulated with 20 mmol / L lactic acid for 24 hours and then irradiated with 75 Gy 6 200 μl of cells / 100 μl of 4T1 cells stimulated with 20 mmol / L lactic acid for 24 hours and then irradiated with 50 Gy.
[0073] Prevention group animal model: select the corresponding experimental mice, randomly di...
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