BeNa Culture Collection
|Culture medium||RPMI-1640 complete medium: 90% RPMI-1640 + 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 it completely 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 3-5 minutes, discard the supernatant, and resuspend the cells with 1-2mL complete culture medium. ③The cell suspension was then added to a T25 flask containing 6-8mL complete medium and placed in an incubator for upright cultivation. Cell subculturing: ① Centrifugation: collect cells, centrifuge at 1000rpm for 5 minutes, discard the supernatant, resuspend the cells with 1-2ml of complete medium, add 1-2mL of culture solution and blow well, and dispense the cell suspension into a fresh T25 flask containing 8ml of culture medium according to a ratio of 1:2. (2) Half-volume solution exchange method: after the culture flask is allowed to stand for 5-10 minutes, half of the supernatant medium is gently sucked away, the remaining medium with cell precipitation is resuspended and mixed evenly, and the cell suspension is dispensed into a fresh T25 flask containing 8ml of medium according to a ratio of 1:2. (3) 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 more than 2 × 106 cells/ml.|
|Growth conditions||37 ℃;5% CO2 95% air;|
|Growth characteristics||Suspension growth|
|Storage conditions||Liquid nitrogen|
|morphology||Lymphoblast-like, round, single cell, clean background, no pigment, no vacuoles|
|Separation substrate||H9 cells are clonal lines (Callo,RC,etal) of HUT78(ATCCTIB161). The cell surface was marked with CD3 and CD4.|
|application||Studies have shown that the cell line is sensitive to human immunodeficiency virus (HIV-1) and can be used to detect, isolate and proliferate HIV-1, as well as to study other human Tcell viruses. Cancer cell line|
|Sharing mode||Public welfare sharing|
H9 human t lymphocyte line
culture :37 ℃,5% CO2 CM2-1 liquid medium
Product format: 2mL frozen vial x 2, or 15mL centrifuge tube 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 culture solution, containing glutamine
(1)prepare a new 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 5-8 days.
Subculture: the suspended cells were gently blowed evenly and distributed into 3-6 fresh culture medium dishes. After being shaken well, the cells were put into the incubator for culture
Cryopreservation: The suspended cells were transferred to the centrifuge tube for (110g, 3min) centrifugation，after centrifugation, terminate digestion with 3ml of cryopreservation solution （90%FBS+10%DMSO), blow evenly and 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|
suspension growth rate ≥ 80.0% in 120 hours
inoculation with 20% cell solution, suspension growth rate reaches 30.0% in 18 hours, and 80.0% in 110 hours
|cell morphology:||suspension, lymphoblast-like||CM2-1 culture medium, suspended, lymphoblast-like, round|
|Conclusion:||good recovery, no abnormal cell morphology, qualified|