human pancreatic stellate cells|BNCC358959 |BNCC

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human pancreatic stellate cells

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 submerse into a 37 ℃ water bath pot, shake the frozen vial to accelerate dissolution, and it is advisable to dissolve all within 1 minute; (2) Add the dissolved cell liquid into a centrifuge tube containing 9ml of complete culture medium in an ultra-clean table, 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, add 1-2mL pancreatin (0.25% Trypsin 0.02% EDTA); (2) Observe the digestion condition under the microscope, when the cell edge shrinks and the adherent is loose (use a pipette to suck up a little pancreatin and gently blow somewhere in the cell layer, the cell layer can be seen to detach with naked eyes, I .e. digestion is completed, otherwise digestion is continued), gently blow the cell layer, blow the cell layer off and blow it away. 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	37 ℃;5% CO2 + 95% air;
Growth characteristics	Adherent growth
Storage conditions	Liquid nitrogen
Type	Adherent growth
Safety level	1
morphology	Fibroblast-like, long fusiform, irregular margin, monolayer adherent growth, clean background
Sharing mode	Public welfare sharing

human pancreatic stellate cell HPaSteC(HPSC)

No. : 358959

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

Validity : 6 years

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% DMEM-H + 10% FBS. DMEM-H:DMEM high sugar culture solution, containing glutamine and sodium pyruvate.

Recovery steps:

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

② take out the frozen storage tube from liquid nitrogen or -80 ℃, take a water bath at 37 ℃ for 1~2min, and move it into the safety cabinet for recovery as soon as possible after it is completely dissolved.

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

④ put it into an incubator (37 ℃,5% CO₂), change the liquid overnight, and fill it in 6-8 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 in about 1min. 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 168 hours	adherence is observed in 18 hours, the cell adherence rate ≥ 80.0% in 150hours
cell morphology:	Adherent, fibroblast-like	Adherent, fibroblast-like, long spindle
attached figure:
conclusion: good viability, no abnormal cell morphology, qualified