Mouse mast cell tumor cells|BNCC339594 |BNCC

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Mouse mast cell tumor cells

Culture medium	RPMI-1640 complete medium: 90% RPMI-1640 + 10% FBS
Subculture procedure	Recovery steps: ① The frozen vial is taken out of liquid nitrogen or -80 ℃ refrigerator and put into PE gloves, quickly submersed into a 37 ℃ water bath pan, shaken The frozen vial to accelerate dissolution, and it is advisable to dissolve all within 1min; (2) Add the dissolved cell solution into a centrifuge tube containing 9ml of complete culture medium in an ultra-clean table, centrifuge at 1000-1200rpm for 5min, discard the supernatant, resuspend the cells with 1-2ml of complete media. ③The cell suspension was then added to the T25 flask containing 6-8mL of complete medium and placed in an incubator for culture. cell subculture: ① transfer the cell suspension solution into a centrifuge tube, centrifuge at 1000rpm for 5min, and collect cells; (2) rinse T25 flask twice with PBS, add 1-2mL pancreatin (0.25% Trypsin + 0.02% EDTA); 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 up some pancreatin, and gently blow somewhere in the cell layer, , the cell layer can be seen to detach with the naked eye, that is, digestion is completed, otherwise digestion continues), directly suck out pancreatin, add 5-6mL of complete medium, gently blow the cell layer off. (3) The cells obtained from the above two steps are evenly mixed and then dispensed into fresh T25 flasks as a ratio of 1:2. add appropriate complete 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	semi-adherent and semi-suspension growth
Storage conditions	Liquid nitrogen
Type	Semi-suspended
Safety level	1
morphology	Epithelial cell-like, short fusiform, round, irregular margin, monolayer adherent growth, clean background
Separation substrate	The cell is derived from a mast cell tumor in DBA/2 mice;
Sharing mode	Public welfare sharing

P815 mouse mast cell

Semi-adherent semi-suspended, epithelial cell-like

No.: 339594

Product format: 2ml frozen vial x 2, or T25 flask x 1

Biosafety level: 1 , handle in ultra-clean table or safety cabinet

Growth Conditions:37 ℃,5% CO₂ CM2-1 culture solution

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,CM2-1 culture solution. CM2-1 culture solution: 90% RPMI-1640 + 10% FBS. RPMI-1640:1640 medium, containing glutamine.

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: The culture medium (including suspended cells) was sucked into the centrifuge tube for centrifugation (110g, 3min) and cells were collected；rinse twice with PBS, add 6ml 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 2minute, and take it out.

(1)Subculture: The digestion of the above dish cells was terminated with 10mL CM2-1 culture medium, and the centrifuge tube cells were suspended with 2mLCM2-1 culture medium. A total of 12mL culture medium was sucked into the dish and mixed evenly, and the culture was divided into 3-6 dishes.

(2)Cryopreservation: Digestion was terminated with 6mL cryopreservation solution (90%FBS+10%DMSO), which was then sucked out and mixed with centrifugal tube cells, divided into 6 cryopreservation tubes, which were cryopreservation at -80℃ with 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 12hours, the cell adherence rate ≥ 80.0% in 70hours	adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 66hours
cell morphology	semi-adherent and semi-suspended, epithelial cell-like	in CM2-1 culture medium, semi-adherent and semi-suspended, epithelial cell-like, short spindle-shaped, round
attached figure
Conclusion	good viability, no abnormal cell morphology, qualified