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HAC15; Adrenal Carcinoma; Human (Homo sapiens) Item ID: ATCCRL3301

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Organism Homo sapiens, human
Tissue adrenal
Product Format frozen
Morphology epithelial-like
Culture Properties adherent
Biosafety Level 1

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.
Disease carcinoma
Age 48 years
Gender female
Ethnicity Black
Applications HAC15 provides an important model system for defining the molecular mechanisms regulating aldosterone and cortisol production and can be applied to studies of either normal adrenal cell function or adrenocortical cancer.
Storage Conditions liquid nitrogen vapor phase
Derivation HAC15 was clonally isolated from NCI-H295R (ATCC® CRL-2128™). NCI-H295R was adapted from the NCI-H295 pluripotent adrenocortical carcinoma cell line (ATCC® CRL-10296™) established by A.F. Gazdar and associates from a carcinoma of the adrenal cortex. The original cells were adapted to a culture medium which decreased the population doubling time from 5 days to 2 days. While the original cells grew in suspension, the adapted cells were selected to grow in a monolayer. The HAC15 cell line responds to ACTH, whereas NCI-H295R lacks this responsiveness.
Genes Expressed aldosterone; cortisol; C19 steroids
Comments HAC15 is a clonal cell line isolated from NCI-H295R (ATCC® CRL-2128™) which, in extended culture, displays a more stable steroidogenic phenotype than NCI-H295R. The HAC15 cells increase aldosterone production in response to treatment with angiotensin II and high extracellular potassium levels. These cells respond to activation cAMP signaling pathway agonists, forskolin and dibutyryl-cAMP, with a time-dependent increase in cortisol and dehydroepiandrosterone production. HAC15 also exhibit a modest increase in cortisol following chronic ACTH (adrenocorticotropic hormone) treatment. In summary, the HAC15 cell line is capable of responding to the three main adrenocortical physiologic regulators.
Complete Growth Medium

To make complete growth medium add to 500 mL DMEM: F12 Medium (ATCC® 30-2006™): 5.6 mL ITS + Premix (Becton Dickenson Cat. No. 354352) and 56 mL Cosmic Calf Serum (Hyclone Cat. No. SH30087.03). 
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. Corning® T-75 flasks (catalog #430641) are recommended for subculturing this product.
  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) (ATCC®30-2200) or 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.
  3. 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.
  4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
  5. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 3 X 104 to 6 X 104 viable cells/cm2 is recommended.
  6. Incubate cultures at 37°C.
Interval: Maintain cultures at a cell concentration between 2 X 105 and 3 X 105 cell/cm2. Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:6 is recommended Medium Renewal: 2 to 3 times per week
Culture Conditions Atmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37°C
STR Profile Amelogenin: X D13S317: 13 D16S539: 11 D5S818: 12 D7S820: 9,12 CSF1PO: 10,12 TH01: 9.3 TPOX: 8 vWA: 17,18
Name of Depositor WE Rainey
References

Wang T, et al. Comparison of aldosterone production among human adrenocortical cell lines. Horm. Metab. Res. 44: 245-250, 2012. PubMed: 22266826

Wang T, et al. Human adrenocortical carcinoma cell lines. Mol. Cell Endocrinol. 351: 58-65, 2012. PubMed: 21924324

Parmar J, et al. Development of an adrenocorticotropin-responsive human adrenocortical carcinoma cell line. J. Clin. Endocrinol. Metab. 93: 4542-4546, 2008. PubMed: 18713819

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