MA-10 (ATCC® CRL-3050)

Organism: Mus musculus  /  Cell Type: Leydig cell tumor  /  Tissue: testes  / 

Organism Mus musculus
Tissue testes
Cell Type Leydig cell tumor
Product Format frozen
Morphology epithelial-like
Culture Properties adherent
Biosafety Level 1
Strain C57BL/6J
Storage Conditions liquid nitrogen vapor phase
Derivation This cell line was derived from transplantable Leydig cell tumor, M5480P, by alternately growing it in culture and in host animals. Cells obtained from culture-derived tumors were cloned.  A clone, designated MA-10, was determined to have retained functional hCG receptors. These cells synthesize progesterone in response to human chorionic gonadotropin (hCG).
Receptor Expression

Luteinizing hormone receptor (LHR)

Comments The MA-10 cell line responds to LH/hCG with increased steroid production. This cell line is a relatively homogeneous cell population. This cell line may provide a suitable model system for the study of the regulation of the expression of differentiated functions of Leydig cells and gonadotropin actions.
Complete Growth Medium The base medium for this cell line is ATCC-formulated DMEM:F12 Medium, Catalog No. 30-2006. To make the complete growth medium, add the following components to the base medium:
  • additional 20mM HEPES
  • horse serum to a final concentration of 15%

Subculturing Note: These cells are cultured on vessels coated with 0.1% Gelatin (ATCC, Cat No. PCS-999-027). 
Note: Use 0.025% Trypsin – 0.053mM EDTA. Dilute 0.25% Trypsin – 0.53mM EDTA (ATCC, Cat. No. 30-2101) 1:10 with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS) (ATCC, Cat. No. 30-2200).
Note: If cell morphology changes from adherent and epithelial-like to being rounded up and forming loosely attached clusters, the serum may not be appropriate for these cells. Change serum lots and/or vendors to obtain correct morphology.  


Volumes used in this protocol are for 75 cm2 flasks; proportionally reduce or increase amount of solutions for culture vessels of other sizes.
1. Remove and discard culture medium.
2. Briefly rinse the cell layers 2 times with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS). 
3. Add 2.0 to 3.0 ml of 0.025% Trypsin-0.053mM 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 gelatin-coated culture vessels. An inoculum of 2 x 104 to 3 x 104 viable cells/cm2 is recommended.
6. Incubate cultures at 37.0°C.
7. Subculture when cultures reach a cell concentration between 2 x 104 to 3 x 105 cells/cm2.


Subcultivation ratio: A subcultivation ratio of 1:3 to 1:8 is recommended.
Medium renewal: 2 to 3 times weekly

Culture Conditions Temperature: 37°C      

Atmosphere: Air, 95%; carbon dioxide (CO2), 5%

Basic Documentation
References

Ascoli M. Characterization of several clonal lines of cultured Leydig tumor cells: gonadotropin receptors and steroidogenic responses. Endocrinology 108(1): 88-95, 1981. PubMed: 6257492