Introduction to cells
In 1957, PuckTT established CHO cells from biopsies of adult Chinese hamster ovaries from CHO-K1, a subclone of CHO. Proline is required for the growth of CHO-K1.
Basic information
CELL ALIAS: CHO CELL CLONE K1;
FOR K1; CHOK1; GM15452
Cell source: Ovary
Cell morphology: epithelial cyto-like
Growth characteristics: adherent growth
Security level: BSL1
Packing: T25 bottles or 1mL cryopreservation tubes
Doubling time: 2~3 days
Passage ratio: 1:3~1:4
Depository: ATCC; CCL-61; ATCC; CRL-9618
CCTCC; GDC0018; JCRB;IFO50414
Culture preservation
Culture system: Ham's F-12K + 10% FBS
Culture conditions:
Gas phase: air, 95%; CO2,5%; Temperature: 37°C
Cryopreservation conditions:
Basal medium + 10% DMSO + 20% FBS, liquid nitrogen preservation
Passaging steps
If the cell density is 80%-90%, it can be subcultured.
- Discard the culture supernatant and rinse the cells with PBS without calcium and magnesium ions 1-2 times;
- Add 2ml of digestion solution (0.25% Trypsin-0.53mM EDTA) to the flask, put it in a 37 °C incubator for 1-2 minutes, and then observe the digestion of the cells under the microscope, if most of the cells become round and fall off, quickly take it back to the operating table, tap the flask a few times and add a small amount of medium to terminate digestion;
- Add medium according to 6-8ml/bottle, gently beat well and aspirate, centrifuge for 4 minutes under 1000RPM conditions, discard the supernatant, add 1-2mL of culture medium and blow well;
- It is recommended that the cell suspension be divided into a new dish or bottle containing 6 ml of medium at a ratio of 1:2 for the first passage after receiving the cells, and it is recommended to freeze one for later use, and subsequent passages are carried out in a ratio of 1:2 to 1:4 according to the actual situation.
Cryopreservation
- When cells are frozen, after discarding the medium, add 1ml of pancreatic enzyme after PBS cleaning, and after the cells become round and detached, add 1ml of serum-containing medium to terminate digestion, and can be counted using a hemocytometer;
- 4min 1000 rpm centrifugation to remove the supernatant. Add 1ml of serum to resuspend cells, add serum and DMSO according to the number of cells, mix gently, the final concentration of DMSO is 10%, the cell density is not less than 1x10 (6)/ml, each cryopreservation tube freezes 1ml of cell suspension, pay attention to the cryopreservation tube to be marked;
- Place the cryopreservation tubes in a programmed cooling box, place them in a -80 degree refrigerator, and transfer to liquid nitrogen irrigation for storage at least 2 hours. Record the location of the cryovials for next retrieval.
Cell use
For scientific use only.
Quality inspection results
Tests for mycoplasma, bacteria, yeast, and fungi are negative.
Notes
1. After receiving the cells at room temperature, please check the packaging status of the product, if you find damage, spillage and contamination, please contact us in time.
2. Do not open the lid of the flask first, treat the surface of the flask with 75% alcohol, if it is adherent cells, please stand in the incubator for 2-3 hours (depending on the cell density status) to stabilize the cell state. If it is a suspension cell, there is no need to stand and follow the suspension cell procedure directly.
3. After standing, confirm the cell growth status under the microscope and take pictures to record. (Photos can effectively help us provide after-sales service)
4. If there is any abnormality in the cell culture process, technical exchanges are welcome.