Human esophageal squamous cell carcinoma cells|359343 |BNCC

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KYSE-150

Subculture procedure	Resuscitation steps: ① Remove the frozen vial from liquid nitrogen or -80 ℃ refrigerator and place it in PE gloves. Quickly immerse it in a 37 ℃ water bath, shake the vial to accelerate dissolution, and dissolve it completely within 1 minute; ② Add the dissolved cell solution into a centrifuge tube containing 9mL of complete medium on a ultra-clean bench, centrifuge at 1000-1200rpm for 5 minutes, discard the supernatant, and resuspend the cells in 1-2mL of complete medium. ③ Add the cell suspension into a T25 flask containing 5-6mL of complete medium and place the flask in an incubator. Cell passage: 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 up some pancreatin and gently blow somewhere in the cell layer, and the cell layer can be seen to detach with naked eyes, I .e. digestion is completed, otherwise digestion is continued), directly suck out pancreatin, add 5-6mL of complete medium, gently blow the cell layer off. ③ Divide the cell suspension into a fresh T25 flask at a ratio of 1:2, add appropriate complete culture medium, mix the cell suspension evenly, and culture it in the incubator.; ④ Pay attention to changes in the pH value of the culture medium and cell density, change the medium regularly (2-3 times a week), and repeat the passage operation or cryopreservation when the cell density reaches 80% -90%.
Growth conditions	37°C; 5% CO₂ + 95% air
Safety level	0
morphology	Epithelial-like，Polygonal，Monolayer adherent growth
Sharing mode	Public welfare sharing

KYSE-150 human esophageal squamous cell carcinoma cells

adherent, epithelial cell-like

No. 359343

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 receiving goods, please contact customer service within 24 hours. if it fails to do so, it will be deemed as good receiving goods. The frozen storage tube shall be stored in a refrigerator at -80 ℃ after receiving the goods. If it is not used for a long time, it shall be transferred to liquid nitrogen storage overnight. T25 form, after receiving the goods, the culture bottle shall be placed in the incubator for rest for 4 hours, and then the cells shall be subjected to routine operation. During recovery, each tube shall not be retained once used up. After recovery, it will be passed on to one generation and can be used normally. Please strictly follow this instruction, otherwise no reissue service will be provided if cell inactivation is caused.

Growth conditions:37 ℃,5% co ₂,90% RPMI-1640/F12 + 10% FBS.

Resuscitation steps:

(1) 1 new 100mm plate containing 12mL of the above culture solution;

(2) the frozen storage tube is taken out of liquid nitrogen or -80 ℃, bathed in water at 37 ℃ for 1~2min, and moved into the safety cabinet for resuscitation as soon as possible after complete dissolution;

(3) using a sterile straw to suck the dissolved liquid into the new plate and shake well clockwise;

(4) put in (37 ℃,5% CO₂) in the incubator, change the liquid overnight and full in 2-4 days.

Subculture/cryopreservation: suck out the old culture solution. after PBS cleaning twice, add 2mL(/100mm dish/T25 bottle) of pancreatin (0.25% Trypsin + 0.02% EDTA) and observe under a microscope. during this period, it is forbidden to shake the culture dish. when the cells just fall off, suck out most of the pancreatin, leave about 0.5mL, move to the incubator for digestion, and take out about 2min. Subculture is terminated by digestion with 6mL of culture solution, and cells are gently blown evenly, and then subculture can be 1:2. For cryopreservation, 3mL of cryopreservation solution (50% basic medium + 40% FBS + 10% DMSO) is used to terminate digestion, blow evenly, divide into 3 frozen storage tubes, and use a program cooling box to freeze storage at -80 ℃.

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 80hours	adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 72hours
cell morphology:	Adherence, epithelial cell-like	adherent, epithelial-like, squamous round
attached figure:
Conclusion:	good viability, and no abnormal cell morphology, qualified

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