Human gastric cancer cells|BNCC337682 |BNCC

Essential Information Certificate Related Products

MKN-45

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) After gently rinsing 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 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 is continued), directly suck out pancreatin, add 5-6mL of complete culture medium, gently blow the cell layer off. (3) The cells obtained from the above two steps are evenly mixed together and then dispensed into fresh T25 bottles according to a ratio of 1:2, appropriate complete culture medium is added, the cell suspension is evenly mixed and cultured in an incubator. ④ Pay attention to the change of pH value of media and cell density, change 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
Safety level	1
morphology	Epithelial cell-like, short fusiform, rounded, irregular margin, clean background
Sharing mode	Public welfare sharing

MKN-45 human gastric cancer cells

semi-adherent semi-suspended, epithelial cell-like

number: 337682

culture:37 ℃,5% CO₂ CM2-1 liquid medium

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.

culture 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 fresh 100mm culture dish containing 12ml of the above culture medium;
(2)  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.
(3)  draw the solution with a sterile pipette, drip into a new culture dish, and mix it evenly by shaking clockwise;
(4) Put it into incubator (37℃，5%CO₂）, change the media overnight, and it will grow up in 5-6 days.

subculture/ cryopreservation: centrifuge the suspended cells at 110g for 3 minutes to collect the cells, rinse twice with PBS, add 2ml of 0.25%Trypsin+0.02%EDTA. Observe under the microscope, remove most of the trypsin(with 0.5ml left) when the cells detach. transfer it to the incubator for digestion about 1.5 minutes, and take it out.

(1) Passage: terminate digestion of cells in culture dish with 4ml of CM2-1 culture media, re-suspend the cells in centrifuge tube with 2ml of CM2-1 culture media, mix the 6ml of the solution evenly and dispense into 3~6 culture dishes.

(2) Cryopreservation: terminate digestion with 3ml of cryopreservation solution （90%FBS+10%DMSO), aspirate and mix evenly with the cells in centrifuge tube, 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 168 hours	adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 168 hours
cell morphology	semi-adherent and semi-suspended, epithelial cell-like	in CM2-1 culture medium, semi-adherent and semi-suspended, epithelial cell-like, polygonal, round
attached figure
Conclusion	good viability, no abnormal cell morphology, qualified

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