AKR cells

Product Name:AKR cells

AKR cellsPost processing

The method of cell transportation is to fill the culture medium and seal the bottle mouth after the cells are cultured in a good condition in the culture bottle. After receiving the cells and returning them to their own laboratory, first open the outer packaging and useSpray the entire bottle with 75% alcohol for disinfection and place it in a clean bench for strict aseptic operation，Incubator standing still2-4 hoursUnder the microscope observation: not exceedingAt 80% confluence, bottled culture medium can be usedCollect to centrifuge tubeIn the middle, join6ml of culture medium, put in37Incubate in a 5% CO2 incubator at ℃; When the confluence degree exceeds 80%, subculture or cryopreservation should be performed according to the situation. Please refer to the cell culture steps for specific procedures.（If the shipment is a sealed culture bottle, remember to loosen the lid of the bottle when placing it in the incubator for cultivation. After passage, it is recommended to use the original culture medium for one bottle and the self-made culture medium for the other bottle）

AKR mouse esophageal cancer cellsCultivation steps

1Revive cells: will containThe freezing tube of 1mL cell suspension was rapidly shaken and thawed in a 37 ℃ water bath, and 5mL of culture medium was added and mixed evenly. Centrifuge at 1000RPM for 5 minutes, discard the supernatant, and add4-6mLBlow the culture medium evenly. Then add all cell suspensions to the culture bottle and culture overnight (or add the cell suspensions)In a 6cm dish）Cultivate overnight. The next day, change the solution and check the cell density.

IIsubculturingIf the cell density reaches80% -90% can be used for subculture.

a）For adherent cells, the following methods can be used for passaging:

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

2. Add1-2ml digestive solution (02Dissolve 5% Trypsin-0.53mM EDTA in a culture bottle and place it in a 37 ℃ incubator for digestion1-2minThen observe the digestion of cells under a microscope. If most of the cells become round and fall off, quickly take them back to the operating table, lightly tap the culture bottle a few times, and then add5ml or morecontain10% serumTerminate digestion of the culture medium.

3. Gently blow the cells, remove and aspirate, centrifuge at 1000RPM for 8-10 minutes, discard the supernatant, add 1-2mL of culture medium, and blow evenly.

4. Press5-6Add culture medium to ml/bottle and divide the cell suspension into new ones in a ratio of 1:25-6In a new dish or bottle of ml culture medium.

b）TheForsuspensionCells can be passaged using the following methods:

Method 1: Collect cells,Centrifuge at 1000RPM for 8-10 minutes, discard the supernatant, add 1-2ml of culture medium and blow evenly. Divide the cell suspension into new dishes or bottles containing 8ml of culture medium in a ratio of 1:2 to 1:5.

Method 2: You can choose the method of half changing the medium. After discarding half of the culture medium, suspend the remaining cells and press the cell suspension1: Divide into new dishes or bottles containing 8ml of culture medium in a ratio of 2 to 1:3.

PS：If the customer receives2ml tubular cells, after receiving the cells, spray the entire tube with 75% alcohol for disinfection and place it in a clean bench or safety cabinet for strict aseptic operation; Transfer tubular cells to T25 culture bottles or6Cm culture dish, add5Mix about ml of culture medium evenly, place it in the incubator overnight for cultivation, and check the cell density: if the density does not exceed80%, change the medium and continue to culture, passaging or freezing depending on the situation. If the density exceeds80%, can be directly passaged (using the same method as above).

IIICell cryopreservation:（Note: For the method of freezing cells in serum-free cryopreservation solution, please refer to our product number:C7001）

1、Cells grow to cover the culture bottleWhen the area is 80%, discard the culture medium in a 25cm2 culture bottle and wash the cells once with PBS;

2、addTransfer approximately 1ml of digestive solution into a culture bottle and observe under an inverted microscope. After the cells have shrunk and become round, add the culture solution to terminate digestion. Gently blow the cells to detach, then transfer the suspension to a 15ml centrifuge tube and centrifuge at 1000rpm for 5 minutes;

3、Resuspend the cells in an appropriate amount of cryopreservation solution and place them in a cryovial;

4、First, place the cell cryopreservation tube in-20 ℃ for 1.5 hours, then transfer it to -80 ℃