F81 (cat kidney cells)

F81 (cat kidney cells)

Product attributes

Product Introduction

genus domestic cat

Age (gender) unknown

Source of organization unknown

Growth characteristics adherent cells

cell morphology Epithelial like cells

Biosafety level 1

Growth medium RPMI-1640＋10% FBS＋1% P/S

Recommended passage ratio 1:2-1:4

Recommended fluid change frequency 2-3 times per week

Freezing conditions

Freezing solution:55% basic medium+40% FBS+5% DMSO

Temperature: Liquid Nitrogen

culture conditions

Gas phase: air,95%； CO2，5%

temperature37℃

Cell processing:

1) Recovery of frozen cells:

Quickly shake and thaw the cryovial containing 1mL of cell suspension in a 37 ℃ water bath, then add it to a centrifuge tube containing 4-6mL of culture medium and mix well. Centrifuge at 1000RPM for 3-5 minutes, discard the supernatant, and resuspend the cells in the culture medium. Then add the cell suspension to a culture bottle (or dish) containing 6-8ml of culture medium and culture overnight at 37 ℃. The next day, observe cell growth and cell density under a microscope.

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

For the passage of adherent cells, the following methods can be referred to:

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

2. Add 0.25% (w/v) -0.53 mM EDTA to culture bottles (1-2mL for T25 bottles and 2-3mL for T75 bottles), digest at 37 ° C for 1-2 minutes (digestion time can be appropriately extended for difficult to digest cells), 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 3-4ml of culture medium containing 10% FBS to terminate digestion.

3. Gently mix and aspirate, centrifuge at 1000RPM for 3-5 minutes, discard the supernatant, add 1-2mL of culture medium and blow evenly. Divide the cell suspension into new T25 bottles in a ratio of 1:2, add 6-8ml of new culture medium prepared according to the instructions to maintain the growth vitality of the cells, and proceed with subsequent passages in a ratio of 1:2 to 1:5 based on the actual situation.

3) Cell cryopreservation: It is recommended to freeze a batch of cell seeds during the first 3 generations of culture after receiving the cells for subsequent experimental use.

Steps for Cell Passage Culture:

(1) When the cell morphology and growth density under the microscope reach 90%, passage is performed.

(2) Discard the culture medium. Wash 1-2 times with PBS, shake well, and discard.

(3) Add EDTA that covers the bottom of the bottle.

(4) Put the culture bottle into a 37 ℃ incubator for digestion, take it out for about 3 minutes, and observe under a microscope whether there is more than 80% of the cells, shrinkage, rounding, and increased intercellular space. Gently tap the culture bottle to remove the remaining cells, add twice the amount to stop digestion, and blow evenly to avoid excessive digestion.

(5) Suck the cell suspension into a centrifuge tube and centrifuge at 1000rpm for 3-5 minutes.

(6) Discard the supernatant, add culture medium to resuspend the cells, and blow the cells to evenly disperse them.

(7) Discard the cell suspension, select a suitable density and inoculate it into a new culture bottle, replenish the culture medium and shake well, then place it in a CO2 incubator at 37 ℃ for cultivation.

(8) Determine the time for fluid replacement or passage based on the growth status of the cells.

(9) Cell cryopreservation

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Processing after cell reception:

1) After receiving the cells, disinfect the bottle wall with 75% alcohol and place the T25 bottle in a 37 ℃ incubator for about 2-3 hours. If you find any damage to the culture bottle, overflow of liquid, or contamination of the cells, please take photos and contact us promptly.

2) Please confirm the cell status under a 4 or 5X microscope, and take 2-3 photos (10 ×, 20 ×) of the newly received cells, as well as one photo of the appearance of the culture bottle, for retention as a basis for the cell status upon receipt during after-sales service.

3) Adherent cells: Cells are placed in a 37 ℃ incubator for 2-3 hours, and their growth and adherence are observed under a microscope. Some adherent cells may detach and form clusters due to vibration during express delivery. If the growth density of cells observed under the microscope is below 60%, the culture medium in the culture bottle can be removed (if there are unattached cells, they need to be recovered by centrifugation and resuspended into the original culture bottle), and 6-8mL of newly prepared culture medium can be added and placed in the cell culture box for further cultivation. If the cell growth density reaches 70% -80% or more, the cells can be passaged. During the passage process, if cells shed due to transportation vibrations, they need to be recovered by centrifugation.