90%RPMI-1640+10%FBS
90%FBS+10%DMSO
1. Please thaw the cells in T25. Please do not thaw the cells directly to a T75 flask or 10 cm culture dish.
2. The cell confluency needs to be strictly controled. The viability of this cell line is relatively low after thawing. It is recommended to use high-quality FBS for thawing or increase FBS to 20% for thawing. After the cells are subcultured for 2 passages, general culture method can be used for subsequenct culturing.
1.Upon arrival, check all containers for leakage or breakage.
2.Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below -130°C, preferably in liquid nitrogen vapor, until ready for use. Please do not store at -80°C. Storage at -80°C may result in loss of viability.
1.Pipette 6-7 mL of complete medium into a 15 mL centrifuge tube.
2.Gently agitate the vial in a 37°C water bath to thaw. Please keep the O-ring and cap out of the water. Thawing should be rapid (approximately 1-2 minutes).
3.Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by spraying with 70% ethanol or wiping with an alcohol cotton pellet. All of the operations from this point on should be carried out in the hood and under strict aseptic conditions.
4.Transfer the vial contents to the step 1 centrifuge tube containing 6-7 mL complete culture medium. And spin at 1100 rpm for 4 mins at room temp to collect the cells.
5.Resuspend cell pellet with 1ml recommended complete medium. And dispense into a T25 flask (or 6 cm culture dish) containing 4 mL of complete medium. (Note: It is suggested that, prior to the addition of the vial contents, the culture vessel containing the complete growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH 7.0 to 7.6.)
6.Incubate the culture at 37°C in a suitable incubator. A 5% CO2 in air atmosphere is recommended if using the medium described on this product sheet.
1.Addition of fresh medium: gently pipette the cells in the flask and take 20 ul of cells for cell counting; According to the cell counting results, aspirate and discard part of the cell suspension, adjust the cell density to 2x10^5~4.0x10^5 cells/mL.
2.Replace fresh medium when a lot cell debris and the medium turned yellow. Centrifuge at 1100 rpm for 4 minutes to collect cells; Resuspend cells and adjust the cell density to 2x10^5~4.0x10^5 cells/mL.