Instructions for 5637 human bladder cancer cells

The company's products are for scientific research purposes only and cannot be used for clinical diagnosis!

Reference materials(Source Literature):

2、 Cell culture operation

1) Resuscitation of cells: The following cell culture and cryopreservation treatments are for reference only. The specific operating steps are mainly based on the accompanying product manual. Quickly shake and thaw the cryovial containing 1 mL of cell suspension in a 37 ℃ water bath, and mix well with 4 mL of culture medium. Centrifuge at 1000 rpm for 3 minutes, discard the supernatant, add 1-2 mL of culture medium, and blow well. Then add all cell suspensions to a culture bottle containing an appropriate amount of culture medium and culture overnight (or add the cell suspension to a 10 cm dish, add about 8 mL of culture medium, and culture overnight). The next day, change the solution and check the cell density.

2) Cell passage: If the cell density reaches 80% -90%, passage culture can be carried out.

a、 Discard the culture supernatant and rinse the cells 1-2 times with PBS that does not contain calcium or magnesium ions.

b、 Add 1 mL of digestion solution (0.25% Trypsin 0.53mM EDTA) to the culture bottle, allowing the digestion solution to infiltrate all cells. Place the culture bottle in a 37 ℃ incubator for 1-2 minutes (depending on the cell condition), and then observe the cell digestion under a microscope. If most of the cells become round and fall off, quickly take them back to the operating table, tap the culture bottle a few times, and add 2-3 ml of culture medium to terminate digestion. Gently mix well and transfer to a sterile centrifuge tube. Centrifuge at 1000 rpm for 5 minutes, discard the supernatant, add 1-2 mL of culture medium, and blow well.

c、 Divide the cell suspension into a new dish or bottle containing 8 mL of culture medium at a ratio of 1:2, and place it in a culture incubator for cultivation. 3) Cell cryopreservation: When the cells are in good growth condition, cell cryopreservation can be performed. Taking T25 bottles as an example below; a、 Collect cells and cell culture medium, transfer them into sterile centrifuge tubes, centrifuge at 1000 rpm for 4 minutes, discard the supernatant, wash with PBS, discard PBS, and perform cell counting.

b、 Add serum-free cell cryopreservation solution according to the number of cells, make the cell density 5 × 106~1 × 107/mL, gently mix well, and freeze 1mL of cell suspension in each cryopreservation tube, paying attention to labeling the tubes properly. Place the cryovial in a -80 ℃ freezer and transfer it to liquid nitrogen for storage after 24 hours. Record the location of the freezer for future retrieval.

3、 Cultivation precautions

1. After receiving the cells, first observe whether the cell bottle is intact and whether there is any leakage or turbidity in the culture medium. If any of the above phenomena occur, please contact us in a timely manner.

2. Carefully read the cell instructions and understand the relevant information about the cell, such as cell morphology, culture medium used, serum ratio, required cytokines, etc., to ensure consistent cell culture conditions. If there are any problems with the cell due to inconsistent culture conditions, the responsibility shall be borne by the customer.

3. Wipe the surface of the cell vial with 75% alcohol and observe the cell status under a microscope. Due to transportation issues, it is normal for some cells to form fragments due to temperature changes and severe collisions. After observing the cell state, disinfect the bottle wall with 75% alcohol and place the T25 bottle in a 37 ℃ incubator for 2-4 hours.

4. Adherent cells can be digested, suspended cells are directly mixed and collected, centrifuged at 900 rpm to 1000 rpm for 3 minutes, and the supernatant is discarded. Add 5 mL of PBS to resuspend the cells, centrifuge at 900 rpm to 1000 rpm for 3 minutes, resuspend the cells in fresh culture medium, and inoculate them into new culture bottles or dishes for cultivation in an incubator.

5. Please ask the customer to use the same culture medium for cell culture under the same conditions.

6. It is recommended that customers take several photos of the cells in the first 3 days after receiving them, recording the cell status for communication and exchange with our technical department. Due to transportation reasons, some sensitive cells may experience instability. Please contact us promptly to inform us of the specific situation of the cells, so that our technical personnel can track and follow up until the problem is resolved.

7. This cell is for scientific research purposes only.

8. Note: The transport medium (infusion medium) cannot be used to culture cells anymore. Please use a newly prepared medium according to the instructions for cell culture conditions to culture cells. After receiving the cells, it is recommended to subculture them at a ratio of 1:2.

9. Note: 1:2 passage refers to transferring one T25 bottle to two T25 bottles or two 6cm dishes. Not one T25 bottle to two 10cm dishes.

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