| BCRJ Code | 0285 |
| Cell Line | MC3T3-E1 Subclone 14 |
| Species | Mus musculus |
| Vulgar Name | Mouse |
| Tissue | Bone, Calvaria |
| Cell Type | Preosteoblast |
| Morphology | Fibroblast |
| Growth Properties | Adherent |
| Age/Ethinicity | newborn / |
| Derivation | A series of subclones were isolated from the cloned but phenotypically heterogeneous MC3T3-E1 cell line. |
| Applications | These cell lines are good models for studying in vitro osteoblast differentiation, particularly ECM signaling. They have behavior similar to primary calvarial osteoblasts. |
| Tumor Formation: | Yes, in immunodeficient mice (forms bone-like ossicles) |
| Products | collagen |
| Biosafety | 1 |
| Addtional Info | The subclones were selected for high or low osteoblast differentiation and mineralization after growth in medium containing ascorbic acid. |
| Culture Medium | The base medium for this cell line is Alpha Minimum Essential Medium with ribonucleosides, deoxyribonucleosides, 2 mM L-glutamine and 1 mM sodium pyruvate, but without ascorbic acid (GIBCO, Custom Product, Catalog No. A1049001 is a sugestion). 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 flask; 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 PBS without calcium and magnesium to remove all traces of serum that contains trypsin inhibitor. Add 2.0 to 3.0 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. Incubate cultures at 37°C NOTE: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 12 in Culture of Animal Cells, a manual of Basic Technique by R. Ian Freshney, 6th edition, published by Alan R. Liss, N.Y., 2010. |
| Subculturing Medium Renewal | Every 2 to 3 days |
| Subculturing Subcultivation Ratio | 1:6 to 1:8 |
| Culture Conditions | Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37°C |
| Cryopreservation | 95% FBS + 5% DMSO (Dimethyl sulfoxide) |
| Thawing Frozen Cells | SAFETY PRECAUTION:
It is strongly recommended to always wear protective gloves, clothing, and a full-face mask when handling frozen vials. Some vials may leak when submerged in liquid nitrogen, allowing nitrogen to slowly enter the vial. Upon thawing, the conversion of liquid nitrogen back to its gas phase may cause the vial to explode or eject its cap with significant force, creating flying debris.
NOTE: It is important to avoid excessive alkalinity of the medium during cell recovery. To minimize this risk, it is recommended to place the culture vessel containing the growth medium in the incubator for at least 15 minutes before adding the vial contents. This allows the medium to stabilize at its normal pH (7.0 to 7.6). |
| References | Wang D, et al. Isolation and characterization of MC3T3-E1 preosteoblast subclones with distinct in vitro and in vivo differentiation/mineralization potential. J. Bone Miner |
| Depositors | Róber Bachinski |
| ATCC | CRL-2594 |
| Cellosaurus | CVCL_5437 |
