A9 (APRT and HPRT negative derivative of Strain L) (ATCC® CCL-1.4)

Organism: Mus musculus, mouse  /  Tissue: subcutaneous connective tissue; areolar and adipose  / 

Organism Mus musculus, mouse
Tissue subcutaneous connective tissue; areolar and adipose
Product Format frozen
Morphology fibroblast
Culture Properties adherent
Biosafety Level 1
Age 100 days
Gender male
Strain C3H/An
Storage Conditions liquid nitrogen vapor phase
Karyotype modal number = 52; range = 46 to 54; This is a hyperdiploid cell line with 24 to 26 marker chromosomes common to all cells., Nine markers (M1 - M9) are common to L-M(TK-) (see ATCC CCL-1.3) and 16 to 17 are found only in A9. The latter markers include Rb(3.5), Rb(4.15), Rb(5.5), T(8;19), Rb(9.17) and 11 to 12 others., Structurally normal chromosomes 4, 15, 16, 17, 18 and X were absent. Note: Cytogenetic information is based on initial seed stock at ATCC. Cytogenetic instability has been reported in the literature for some cell lines.
Derivation
The A9 cell line was isolated by J.W. Littlefield from wild-type Strain L cells.
Clinical Data
male
Antigen Expression
H-2k
Tumorigenic Yes, Tumors developed within 21 days at 100% frequency (5/5) in nude mice inoculated subcutaneously with 10exp7 cells.
Effects
Yes, in nude mice
Comments
The cells are sensitive to HAT selection medium and deficient in adenosine phosphoribosyl transferase (APRT) and hypoxanthine phosphoribosyl transferase (HPRT).
The cells are resistant to 8-azaguanine (0.003 mg/mL) and 6-thioguanine (0.1 mM).
Complete Growth Medium Dulbecco's modified Eagle's medium with 4.5g/L glucose, 90%; fetal bovine serum, 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.

  1. Remove and discard culture medium.
  2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.03% (w/v) EDTA solution to remove all traces of serum which 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. 
  6. Incubate cultures at 37°C.

Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:10 is recommended

Medium Renewal: Every 2 to 3 days

Cryopreservation
Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO
Storage temperature: liquid nitrogen vapor phase
Culture Conditions
Atmosphere: Air, 95%; carbon dioxide (CO2), 5%
Temperature: 37°C
Name of Depositor TR Chen
References

Littlefield JW. The inosinic acid pyrophosphorylase activity of mouse fibroblasts partially resistant to 8-azaguanine. Proc. Natl. Acad. Sci. USA 50: 568-573, 1963. PubMed: 14067106

Chen TR. Cytogenetics of somatic cell hybrids. II. Modal karyotypes and mosaicism in mouse mutant cell lines: LM(tk-), A9, and Rag. Cancer Genet. Cytogenet. 24: 95-107, 1987. PubMed: 3466673

Mayr GA, Freimuth P. A single locus on human chromosome 21 directs the expression of a receptor for adenovirus type 2 in mouse A9 cells. J. Virol. 71: 412-418, 1997. PubMed: 8985365

Basic Documentation
References

Littlefield JW. The inosinic acid pyrophosphorylase activity of mouse fibroblasts partially resistant to 8-azaguanine. Proc. Natl. Acad. Sci. USA 50: 568-573, 1963. PubMed: 14067106

Chen TR. Cytogenetics of somatic cell hybrids. II. Modal karyotypes and mosaicism in mouse mutant cell lines: LM(tk-), A9, and Rag. Cancer Genet. Cytogenet. 24: 95-107, 1987. PubMed: 3466673

Mayr GA, Freimuth P. A single locus on human chromosome 21 directs the expression of a receptor for adenovirus type 2 in mouse A9 cells. J. Virol. 71: 412-418, 1997. PubMed: 8985365