May serve as a platform for the evaluation of several chemotherapeutic strategies to treat chordoma as it accurately recapitulates chordoma. The Brachyury expression may facilitate understanding of its role in other epithelial-derived cancers.

The Chordoma Foundation may be able to offer financial assistance for the purchase of this cell line. Please contact cells@chordoma.org for more information.

Fresh surgical chordoma tissue was minced in 0.25% trypsin then placed on shaker at 37°C for 30 minutes. The tissue was then triturated and passed through 40 µm nylon mesh. Single cells were plated and the culture expanded. (source: PubMed: 21699479)

CRL-3267 is a primary chordoma cell line from a sacral tumor that expresses Brachyury. This cell line’s classical phenotypic features, such as characteristic physaliferous cells and stable expression of Brachyury, are maintained through serial passaging.

Expresses embryonic transcription factor Brachyury. This cell line was accessioned with the support of the Chordoma Foundation, a non-profit organization working to improve the lives of chordoma patients by accelerating research to develop effective treatments for the chordoma disease.

Volumes used in this protocol are for 75 cm² flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

1. Remove and discard culture medium. Briefly rinse the cell layer with Ca⁺⁺/Mg⁺⁺ free Dulbeccos phosphate-buffered saline (D-PBS) (ATCC 30-2200) or 0.25% (w/v) Trypsin - 0.53 mM EDTA (ATCC 30-2101) solution to remove all traces of serum which contains trypsin inhibitor.
2. 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.
3. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.
4. Transfer cell suspension to a centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes.
5. Discard supernatant. Resuspend the cell pellet in fresh growth medium.
6. Add appropriate aliquots of the cell suspension to new culture vessels
7. Incubate cultures at 37°C.

xa0

Subcultivation Ratio: 1:2 to 1:3 is recommended.

Medium Renewal: 2 to 3 times a week

Hsu W, et.al. Generation of chordoma cell line JHC7 and the identification of Brachyury as a novel molecular target. J. Neurosurg. 115(4): 760-769, 2011. PubMed: 21699479