HBL human melanoma cells

HBL human melanoma cells

Product Name:HBL human melanoma cells

Brand: Shanghai Yanzun Biotechnology
Delivery time: In stock
Cell count: 1x10 ^ 6 cells
Cell viability: 95% (Viability by Trypan Blue Exclusion)
Cell passage: 1:4~1:6 passage; Change the fluid 2-3 times a week
Growth conditions: Gas phase: Air, 95%; Carbon dioxide, 5%; Temperature: 37 ℃
Cell detection: Cells do not contain HIV-1, HBV, HCV, mycoplasma, bacteria, yeast, and fungi
Cultivation conditions: DMEM (high sugar)+10% FBS+1% P/S; 37℃，5% CO2
Passage method: It is recommended to change the medium every two days at a ratio of 1:2-1:3
Freezing conditions: serum-free cell freezing solution
Freezing conditions: 90% FBS+10% DMSO
Cell cryopreservation: liquid nitrogen cryopreservation

Cell culture operation
Dry ice transportation: After receiving the cells, they need to be immediately transferred to liquid nitrogen storage, or frozen in a -80 ° C freezer for a short period of time, or directly revived
Room temperature transportation: After receiving the cells, please immediately remove the cell culture bottle from the packaging box and follow the steps below for cultivation and passage
Cell passage: When the cell density reaches 80% -90%, passage culture can be carried out
Steps for operating cells in culture bottles
For adherent cultured cells, we will fill the entire culture bottle with culture medium before shipping to reduce the shedding of adherent cells during product transportation.
After receiving the cells, please pay attention to whether there is contamination. Place the culture bottle under an inverted microscope and carefully inspect for turbidity and bacterial contamination. Due to the bumps during transportation and the sensitivity of some cells to temperature changes, there may be some cells shedding and floating. These cells are still alive and should not be discarded. They can be centrifuged and enriched before use.
2. For adherent cells, in a biosafety cabinet environment, use a vacuum pump to remove excess culture medium from the culture bottle until about 5-8mL remains. Then, select the appropriate CO2 content and culture in a 37 ℃ constant temperature incubator according to the culture medium, and loosen the bottle cap. If the cells have already filled the culture bottle, please subculture immediately.
3. For suspended cells, transfer the cells from the culture bottle to a centrifuge tube in a biosafety cabinet environment, centrifuge at 150 g for 5 minutes, remove the supernatant, blow off the cells with 5 mL of culture medium, transfer to a new culture bottle, and then incubate in a 37 ℃ constant temperature incubator with appropriate CO2 content.
Operation steps for freezing tube cells
Attention: To ensure high cell viability, please thaw and culture immediately after receiving the product.
1. Place the cryovial in a 37 ℃ water bath and shake it back and forth to quickly thaw. To avoid pollution, ensure that the frozen storage pipe is placed above the water surface. Thawing should be done quickly, about 1 minute.
Once the liquid in the cryovial melts, immediately remove it and use alcohol to spray the surface of the cryovial. Starting from this step, subsequent operations must be completed in the biosafety cabinet.
3. Transfer the liquid from the cryovial to a centrifuge tube containing 5mL of complete culture medium, centrifuge at 150g for 5 minutes, and remove the supernatant using a vacuum pump.
4. After using the culture medium, resuspend the cells and transfer them to a new culture bottle. To ensure the survival rate of cell recovery, please preheat the culture medium in a 37 ℃ water bath before use.
5. Place the cells in a 37 ℃ constant temperature incubator containing appropriate CO2 for cultivation.
Subculture of adherent cells
1. Suck and discard the culture medium from the culture bottle, add PBS and rinse once.
2. Add 1 mL of 0.25 (w/v) Trypsin EDTA solution and incubate in a 37 ℃ incubator until the cells detach from the wall. This process takes approximately 3-5 minutes.
3. Add 2mL of culture medium to neutralize trypsin, gently blow the cells off the surface of the culture bottle, and disperse the cells.
4. Collect the cell suspension into a 15mL centrifuge tube, centrifuge at 150g for 5 minutes, and remove the supernatant using a vacuum pump. Take an appropriate amount of culture medium to resuspend the cells and transfer them to a new culture bottle for cultivation.
Suspension cell passaging can refer to the following methods:
1、 Direct passage method
Once the suspended cells have grown to 80% to 90% (the cell suspension has turned yellow), they can be passaged;
2. Use a straw to aspirate 1/2~1/3 of the cell suspension;
3. Add an appropriate amount of fresh culture medium and continue cultivation.
2、 Centrifugal passaging method (in the presence of cell debris)
1. Transfer the cell suspension into a centrifuge tube;
Centrifuge at 2.150 g for 5 minutes and discard the upper clear liquid;
3. Resuspend cells using fresh culture medium;
4. Suck an appropriate amount of cell suspension with a straw, place it in a new culture bottle, add an appropriate amount of fresh culture medium, and continue culturing.
Cell cryopreservation operation
Centrifuge at 1000 rpm for 5 minutes to remove the supernatant. Add 1 mL of serum to resuspend cells, add serum and DMSO according to the number of cells, gently mix well, DMSO final concentration is 10%, cell density is not less than 1 x 106/mL, each cryovial is frozen with 1mL of cell suspension, pay attention to labeling the cryovials;
Place the cryotube in a program cooling box, put it in a -80 ° C refrigerator, and after at least 2 hours, transfer it to liquid nitrogen storage

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