Human nephroblastoma cell|BNCC352207 |BNCC

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Human nephroblastoma cell

Culture medium	McCoy's 5a complete medium (CM24-1):85% McCoy's 5a + 15% 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 6-8mL of complete medium and placed in an incubator for culture. Cell subculturing: ① transfer the cell suspension solution into a centrifuge tube, centrifuge at 1000rpm for 5 minutes to collect cells; (2) rinse the T25 flask twice with PBS, add 1-2mL pancreatin (0.25% Trypsin + 0.02% EDTA); Observe the digestion situation under the microscope. When the edge of the cell shrinks and the adherent is loose (a pasteur pipette can be used to suck up some pancreatin, 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 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;
Storage conditions	Liquid nitrogen
Type	Semi-adherent growth
Safety level	1
morphology	In the CM5-1 culture solution, semi-adherent, irregular, round and short shuttle
Separation substrate	renal, pleural exudate
application	The microstructure has a few microvilli, connecting complexes, and a complete Golgi apparatus. The endoplasmic reticulum is mostly smooth, lipid droplets, and no virus particles.
Sharing mode	Public welfare sharing

SK-NEP-1 human nephroblastoma cells

Semi-adherent, irregular cells

No. : 352207

Growth :37 ℃,5% CO₂ CM24-1 culture solution

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

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,CM24-1 culture solution. CM5-1 culture solution: 85% McCoy & rsquo; S 5a + 15% FBS. McCoy‘s 5a:MC，5A 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 3-5 days.

Subculture/ cryopreservation: remove old medium, and rinse twice with PBS,add 12ml culture media directly,blow the cells with a pipette until they fall off completely, aspirate and dispense into 3~6 culture dishes;Cryopreservation: The suspended cells were transferred to the centrifuge tube for (110g, 3min) centrifugation，after centrifugationterminate 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 12 hours, the cell adherence rate ≥ 80.0% in 86 hours	adherence is observed in 18 hours, cell adherence rate ≥ 80.0% in 96 hours
cell morphology	semi-adherent, irregular cell	in the CM24-1 culture solution, it is semi-adherent, irregular, round and short shuttle
attached figure
conclusion	good viability, and no abnormal cell morphology, qualified

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