IDH1 mutant-U-87 Isogenic Cell Line (ATCC® HTB-14IG)

Organism: Homo sapiens, human  /  Tissue: brain  /  Disease: glioma

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Organism Homo sapiens, human
Tissue brain
Product Format frozen 1.0 mL
Morphology epithelial
Culture Properties adherent
Biosafety Level 2  [Cells containing CMV and SV40 viral sequences]

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 glioma
Gender male
Ethnicity unknown
Applications Isocitrate dehydrogenase 1; IDH1 mutant; IDH1R132H mutation, anti-cancer drug screening, tumor-associated IDH1 mutation, IDH1-specific inhibitors.
Storage Conditions liquid nitrogen vapor phase
Images ATCC HTB-14IG Bio-functional validation data ATCC HTB-14IG, IDH1 mutant-U-87 Isogenic Cell Micrograph
Comments

This is a glioma IDH1R132H mutant isogenic line derived from the parental U-87MG (ATCC® HTB-14) cell line. The c.395G>A knock-in mutation encoding IDH1R132H protein expression was generated at ATCC by utilizing the CRISPR/Cas9 gene editing technology. This is a heterozygous mutation expressing the c.395G>A mutant allele. The IDH1R132H mutation in HTB-14IG has been validated at the genomic, transcript, and protein bio-functional levels.

The IDH1R132H mutation is among the most common seen in glioma and serves as an important diagnostic marker in various stages of disease in patients. This IDH1R132H mutant isogenic line HTB-14IG has been tested at ATCC for neomorphic functional activity displaying elevated levels of intra- and extracellular D-2HG above the parental U-87MG line. In addition, the HTB-14IG IDH1 mutant isogenic line displays histone hypermethylation compared to the parental line. This IDH1R132H isogenic cell model is a valuable in vitro cell-based tool for clinical diagnostics, elucidating mechanisms involved in cancer-associated differentiation, tumorigenesis and use in screening anti-cancer compounds for drug discovery and development.

Refer to the parental line U-87MG (ATCC® HTB-14) for additional background information.

Complete Growth Medium

The base medium for this cell line is Eagle's Minimal Essential Medium (EMEM; ATCC 30-2003). To make the complete medium add Fetal Bovine Serum (FBS; ATCC 30-2020) to the base medium for 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. 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 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.
    Cultures can be established between 2.0 x 104 and 6 x 104 viable cells/cm2.
  6. Incubate cultures at 37°C.
Interval: Maintain cultures at a cell concentration between 2 X 104 and 1 X 105 cell/cm2.
Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:6 is recommended
Medium Renewal: 2 to 3 times per week
Cryopreservation Complete growth medium supplemented with 5% (v/v) DMSO (ATCC 4-X)
Culture Conditions
Atmosphere: air, 95%; carbon dioxide (CO2), 4 to 6%
Temperature: 36.0 to 38.0°C
Cells per Vial ≥ 1 x 106 cells
Volume 1.0 mL
STR Profile
Amelogenin: X
CSF1PO: 10,11
D13S317: 8,11
D16S539: 12
D5S818: 11,12
D7S820: 8,9
THO1: 9.3
TPOX: 8
vWA: 15,17
Sterility Tests Bacteria and yeast: No growth
Mycoplasma: No growth
Viral Testing Hepatitis B: None detected
Cytomegalovirus: None detected
Human immunodeficiency virus: None detected
Epstein-Barr virus: None detected
Human papillomavirus: None detected
Functional Tests Genotype testing for knock-in mutation - heterozygous DNA change: c.395G>A mutation (correlates to protein sequence p.R132H).
Elevated intra and extracellular level of D-2-hydroxyglutarate (D-2HG) measurement and histone hypermethylation compared to parental line confirmed during product development.
Population Doubling Time 25.6 hours
Year of Origin 2016
References

Avellaneda Matteo D, et al. Molecular mechanisms of isocitrate dehydrogenase 1 (IDH1) mutations identified in tumors: The role of size and hydrophobicity at residue 132 on catalytic efficiency. J Biol Chem 292(19): 7971-7983, 2017. PubMed: 28330869

Bralten LB, et al. IDH1 R132H decreases proliferation of glioma cell lines in vitro and in vivo. Ann Neurol 69(3):455-463, 2011. PubMed: 21446021

Rossetto M, et al. Metabolism of glioma and IDH1/IDH2 mutations. Rev Neurol 167(10): 699-703, 2011. PubMed: 21885076

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation
Other Documentation
FAQ's
  1. Validation - HTB-14IG


    Date Updated: 10/3/2017

  2. IDH1 R132H human glioma isogenic cell line


    Date Updated: 10/3/2017

  3. Applications of the IDH1 mutant U-87 Isogenic Cell Line


    Date Updated: 10/3/2017

Restrictions

For commercial accounts, this cell line is only distributed under the terms of a fully signed and executed ATCC® Material Transfer Agreement and Addendum.  If the commercial account is screening per completed Addendum, the recipient will be required to pay a Screening Fee (ATCC® ACS-2103).

Screening Use is defined as use of Biological Material in small molecule and biologic drug discovery, including initial target identification and validation, assay development, high throughput screening, hit identification, lead optimization, and selection of candidates for clinical development.

If the commercial account is not screening per the completed Addendum, the recipient will not be required to pay a Screening Fee.

In addition to the foregoing, this product's use is governed by the CRISPR Label License Agreement. For information on purchasing a license to use this product for purposes other than those permitted in the CRISPR Label License Agreement, please contact The Broad Institute at partnering@broadinstitute.org.

References

Avellaneda Matteo D, et al. Molecular mechanisms of isocitrate dehydrogenase 1 (IDH1) mutations identified in tumors: The role of size and hydrophobicity at residue 132 on catalytic efficiency. J Biol Chem 292(19): 7971-7983, 2017. PubMed: 28330869

Bralten LB, et al. IDH1 R132H decreases proliferation of glioma cell lines in vitro and in vivo. Ann Neurol 69(3):455-463, 2011. PubMed: 21446021

Rossetto M, et al. Metabolism of glioma and IDH1/IDH2 mutations. Rev Neurol 167(10): 699-703, 2011. PubMed: 21885076