BeNa Culture Collection
|Culture medium||DMEM-H complete medium: 90% DMEM-H + 10% FBS|
|Subculture procedure||Recovery steps: ① Take out the frozen vial from liquid nitrogen or -80 ℃ and put it into PE gloves, quickly subtract into a water bath pot at 37 ℃, shake the frozen vial to accelerate dissolution, and completely dissolve within 1 minute. (2) Add the dissolved cell solution to 9ml in the ultra-clean table; In the centrifuge tube of complete culture medium, centrifuge at 1000-1200rpm/min for 5 minutes, discard the supernatant, resuspend the cells with 1-2ml of complete media. ③The cell suspension was then added to the T25 flask containing 5-6mL of complete medium and placed in an incubator for cultivation. Cell subculturing: ① remove the medium, rinse twice with PBS, and add 1-2mL pancreatin (0.25% Trypsin + 0.02% EDTA); (2) Observe the digestion situation under the microscope. When the cell edge shrinks and the adherent is loose (a pasteur pipette can be used to suck out a little pancreatin and gently blow somewhere in the cell layer, the cell layer can be seen to detach with naked eyes, that is, digestion is completed, otherwise digestion is continued), directly suck out pancreatin add 5-6ml of complete medium, gently blow the cell layer off. ③ Dispense the cell suspension into a fresh T25 flask as the ratio of 1:2, add appropriate complete culture medium, mix the cell suspension evenly, and culture in an incubator. ④ Pay attention to the change of pH of medium and cell density, renew the medium regularly (2-3 times a week), and repeat the subculture or cryopreservation when the cell density reaches 80%-90%.|
|Growth conditions||Culture temperature 37 ℃; Atmosphere 5% CO2 + 95% air;|
|Growth characteristics||Adherent growth|
|Storage conditions||Liquid nitrogen|
|morphology||Fibroblast-like, long spindle|
|application||This is one of the cell lines constructed by J. Ponten and his colleagues from malignant glioma between 1966 and 1969 (others include ATCC HTB-14 and ATCC HTB-16 and ATCC HTB-17). In 1987, mycoplasma contamination was removed by BM-Cycline culture for 6 weeks.|
|Sharing mode||Public welfare sharing|
U-118 MG human astroblastoma
Adherence, diverse forms
Product format: 2ml frozen vial x 2, or T25 flask x 1
Biosafety level: 1 , handle in ultra-clean table or safety cabinet
Receiving notice: if any abnormality is found on the day of receipt, please contact the customer service within 24 hours. If it is overdue, it is deemed that the cells are well. For the frozen cells, it shall be stored in the refrigerator at -80 ℃ upon arrival. If they are not used for a long time, they shall be transferred to liquid nitrogen for storage overnight. Recovered cells in T25 culture flask, upon receipt, put the culture flask in the incubator for 4h, and then carry out handling procedure. During recovery, each vial shall be used up once and shall not be retained. After recovery, the cells can be passed on to the next generation and can be used normally. Please operate in strict accordance with this instruction, otherwise the replacement of cells are not be available in case of lose of cell viability.
Growth conditions :37 ℃,5% CO2,CM1-1 culture solution. CM1-1 culture solution: 90% DMEM-H + 10% FBS. DMEM-H:DMEM high sugar culture solution, containing glutamine and sodium pyruvate.
(1)Prepare a new 100mm culture dish containing 12ml of the above culture medium;
remove the frozen vial from liquid nitrogen or refrigerator at -80℃, thaw the vial in 37°C water bath for 1-2 minutes. Transfer the vial to biosafety cabinet for culture as soon as the contents are completely thawed.
(2)Draw the solution with a sterile pipette, drip into a new culture dish, and mix it evenly by shaking clockwise;
(3)Put it into incubator (37℃，5%CO2）, change the media overnight, and it will grow up in 4-5 days.
Subculture/ cryopreservation: remove old medium, and rinse twice with PBS, add 2ml of 0.25%Trypsin+0.02%EDTA, do not shake the culture dish during the period. Observed under the microscope, remove most of the trypsin(with 0.5ml left) when the cells detach. transfer it to the incubator for digestion about 1minute, and take it out.
(1) Subculture: terminate digestion with 6ml of CM1-1 culture media, aspirate and dispense into 3~6 culture dishes.
(2) Cryopreservation: terminate digestion with 3ml of cryopreservation solution （90%FBS+10%DMSO), aspirate and dispense into 3 frozen vials, and cryopreserve at -80 ℃ with a programmed cooling box.
Recovery record: According to the recovery instructions, the results of the cell recovery are reported as follows:
|item||quality standard||recovery record|
|viability||adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 120hours||adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 108hours|
|cell morphology||adherent, diverse forms||in the CM1-1 culture solution, it is adherent, mixed in form, low density is epithelial cell-like, high density is fibroblast-like, polygonal, long spindle|
|conclusion||good viability, and no abnormal cell morphology, qualified|