Human neuroblastoma cells|BNCC337645 |BNCC

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Human neuroblastoma cells

Culture medium	DMEM-H/F12 complete medium: 90% DMEM-H/F12 + 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 culture. 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
Safety level	1
morphology	Epithelial cell-like, short fusiform, polygonal, rounded, irregular margin, monolayer adherent growth, clean background, no pigment, no vacuoles
Sharing mode	Public welfare sharing

SK-N-BE(2) human neuroblastoma cells

adherent, epithelial cell-like

No.: 337645

Culture:37 ℃,5% CO₂ CM9-1 culture solution

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 loss of cell viability.

Growth conditions:37 ℃,5% CO₂,CM9-1 culture solution. CM9-1 culture solution: 90% DMEM-H/F12 + 10% FBS. DMEM-H/F12:DMEM high sugar culture solution is mixed with F12 culture solution 1:1, containing glutamine and sodium pyruvate.

Recovery steps:
(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 2-4 days.

Subculture/ cryopreservation: remove old medium, and rinse twice with PBS, add 6ml trypsin, 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 3minute, and take it out.

(1) Subculture: terminate digestion with 12ml of CM9-1 culture media, aspirate and dispense into 3~6 culture dishes.

(2) Cryopreservation: terminate digestion with 6ml of cryopreservation solution （90%FBS+10%DMSO), aspirate and dispense into 6 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 70hours	adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 66hours
cell morphology:	adherent, epithelial cell-like	in CM9-1 culture solution, adherent, epithelial cell-like, spindle-shaped, polygonal, a few round
attached figure:
Conclusion:	good viability, no abnormal cell morphology, qualified

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