Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Age
adult
Applications
This cell line is insensitive to epsilon toxin from C.perfringens and is a control for MDCK.2 (ATCC CRL-2936)
Storage Conditions
liquid nitrogen vapor phase
Karyotype
hyperdiploid canine cell line with a modal chromosome number of 76 with low polyploidy rate. Several unidentifiable marker chromosomes were present in most of the cells examined.
Images
Derivation
Cell line was derived by cloning (limited dilution) the parental cell line MDCK (ATCC CCL-34).
The base medium for this cell line is ATCC-formulated Eagle's Minimum Essential Medium, Catalog No. 30-2003. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.
Subculturing
Volumes used in this protocol are for 75 cm2 flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
Remove and discard culture medium.
Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) or 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.
Add 1.0 to 2.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes). Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 1 X 104 to 3 X 104 viable cells/cm2 is recommended.
Incubate cultures at 37°C. Subculture when the cell concentration is between 8 X 104 and 2 X 105 cells/cm2.
Subcultivation ratio: A subcultivation ratio of 1: 3 to 1:8 is recommended.
