Pharmaceutical composition and uses thereof
A composition and drug technology, applied in drug combinations, antitumor drugs, pharmaceutical formulations, etc., can solve problems such as unsatisfactory, rare, and negative effects, and achieve the prevention of tumor occurrence, less toxic and side effects, and a wide antigen spectrum. Effect
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Embodiment 1
[0038] Example 1 Cell Culture
[0039] Experimental materials: H22 cell line, 10% fetal bovine serum, penicillin and streptomycin mixture (Beijing Tigemei Company).
[0040] Experimental method: culture H22 cell line in the culture medium containing 10% fetal bovine serum, penicillin 100IU / mL, streptomycin 100×μg / mL, at 37°C and 5% CO 2 Culture in the incubator, when the cells grow to the logarithmic phase, press 3×10 6 / 100ml inoculation.
Embodiment 2
[0041] Embodiment 2 drug treatment
[0042] Experimental materials: procaine, MS-275 (both purchased from sigma company).
[0043] Experimental grouping: the volume of cell culture medium is strictly consistent in each group, blank control group, exosomes control group (no drug addition group), exosomes experimental group 1 (adding procaine), exosomes experimental group 2 (adding drug MS-275) , exosomes experimental group 3 (combined administration of procaine and MS-275).
[0044] Experimental method: in the control group, no drug was added after inoculation, and 150ml of the supernatant was collected as a control after 24 hours; -6 mol / L of procaine, respectively collected 150ml of culture supernatant 24h after dosing; dosing group 2, 24h after inoculation with a concentration of 1 × 10 -6 mol / L MS-275 treatment, respectively collected 150ml of culture supernatant 24h after dosing; dosing group 3, 24h after inoculation with 1 × 10 -6 mol / L procaine and 1×10 -6 mol / L MS-2...
Embodiment 3
[0045] The separation and purification of embodiment 3 exosomes
[0046] Experimental equipment: HMAC-CP70G low temperature ultra-high speed centrifuge, 100KU MW CO Millipore Amicon high recovery rate high flow rate tangential flow ultrafiltration centrifuge tube (Millipore Company).
[0047] Experimental method: Centrifuge the collected cell culture supernatants of the experimental group and the control group at 300g for 10min to remove the cells, and take the supernatant; centrifuge at 1500g for 30min to remove the cell debris, collect the supernatant, and concentrate it through a 100kUMWCO Centriplus centrifugal ultrafiltration tube Ultrafiltration, centrifugation at 1500g for 30min to obtain 6ml of concentrated solution, transfer the separated and purified concentrated solution to a 1.5ml centrifuge tube, and ultracentrifuge at 100kg for 60min at 4°C with a horizontal rotation angle to obtain exosomes.
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