I-13.35 (ATCC® CRL-2471)

Organism: Mus musculus, mouse  /  Cell Type: macrophage  /  Tissue: spleen  / 

Organism Mus musculus, mouse
Tissue spleen
Cell Type macrophage
Product Format frozen
Morphology macrophage
Culture Properties adherent
Biosafety Level 1
Age adult
Gender female
Strain C3H/HeJ
Applications

The cell line can be used to phenotype and characterize the role of splenic macrophages in the initiation of immune responses.

Storage Conditions liquid nitrogen vapor phase
Derivation

This is an immortalized cell line derived from the progeny of individual splenic macrophage progenitor cells of an apparently normal adult mouse. Cells were cloned in soft agar in the presence of CSF-1 and expanded on microcarrier beads. Beads were transferred to culture dishes and were subsequently passaged by scraping.

 

Antigen Expression
CD11b/CD18 (Mac-1) +, MHC Class I +, MHC Class II +, CD115 (colony stimulating factor 1 receptor (CSF-1R)) +
Receptor Expression
colony stimulating factor 1 (CSF-1R, CD115)
Comments The cell line possesses many features of mature macrophages, including antibody-dependent phagocytic and cellular cytotoxic activities, ability to secrete lysozyme, and expression of the Mac-1 antigen and mRNA for the CSF-1 receptor. RefWilson CM, et al. Immortalization of growth factor-dependent mouse splenic macrophages derived from cloned progenitors. J. Immunol. Methods 137: 17-25, 1991. PubMed: 1707081

This cell line, unlike I-11.15 (ATCC CRL-2470), has the constitutive ability to present antigen to Th1 helper T cells and it expresses higher levels of MHC Class II antigens than I-11-15. RefMcCormack JM, et al. Mouse splenic macrophage cell lines with different antigen-presenting activities for CD4+ helper T cell subsets and allogeneic CD8+ T cells. Cell. Immunol. 145: 359-371, 1992. PubMed: 1451184

C3H/HeJ strain is defective in TLR4 (toll-like receptor 4)

Complete Growth Medium Dulbecco's modified Eagle's medium with 4 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate and 4.5 g/L glucose, 70%; fetal bovine serum, 10%; LADMAC Conditioned Media (produced from the LADMAC cell line (CRL-2420), 20%
Subculturing
  1. Remove and discard 75% of culture medium.
  2. Scrape cells with cell scraper.
  3. Add appropriate aliquots of cell suspension to new culture vessels.
  4. Place culture vessels in incubators at 37°C.

Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:4 is recommended
Medium Renewal: Every 2 to 3 days

LADMAC Conditioned Medium

LADMAC conditioned medium is made from LADMAC cells (ATCC CRL-2420).  LADMAC cells secrete the growth factor colony stimulating factor 1 (CSF-1). CSF-1 is capable of supporting the in vitro proliferation of mouse bone marrow macrophages. The Pannell-Milstein roller bottle apparatus may be used to produce high concentrations of CSF-1 RefOlivas E, et al. Use of the Pannell-Milstein roller bottle apparatus to produce high concentrations of the CSF-1, the mouse macrophage growth factor. J. Immunol. Methods 182: 73-79, 1995. PubMed: 7769247

Subculturing Procedure for LADMAC cells

Cultures can be established by centrifugation with subsequent resuspension at 2 X 105 viable cells/mL. Attached cells may be dispersed by tapping the sides of the flask until cells are loose.

Complete Growth Medium for LADMAC cells

The base medium for this cell line is ATCC-formulated Eagle's Minimum Essential Medium, Catalog No. 30-2003.  To make the complete growth medium, add the following components to the base medium:  fetal bovine serum to a final concentration of 10%

This medium is formulated for use with a 5% CO2 in air atmosphere.

Conditioned medium harvesting:

  1. Allow LADMAC cells to become confluent.
  2. After 5 to 7 days, collect supernate, centrifuge at 125 x g for 5 to 10 minutes
  3. Filter (200 nM filter) and tore aliquots at –20oC
Cryopreservation
Culture medium 95%; DMSO, 5%. Cell culture tested DMSO is available as ATCC Catalog No. 4-X.
Culture Conditions
Temperature: 37°C
Name of Depositor WS Walker
Passage History
Cells were cloned in soft agar in the presence of CSF-1 and expanded on microcarrier beads. Beads were transferred to culture dishes and were subsequently passaged by scraping.
References

Olivas E, et al. Use of the Pannell-Milstein roller bottle apparatus to produce high concentrations of the CSF-1, the mouse macrophage growth factor. J. Immunol. Methods 182: 73-79, 1995. PubMed: 7769247

Wilson CM, et al. Immortalization of growth factor-dependent mouse splenic macrophages derived from cloned progenitors. J. Immunol. Methods 137: 17-25, 1991. PubMed: 1707081

Basic Documentation
References

Olivas E, et al. Use of the Pannell-Milstein roller bottle apparatus to produce high concentrations of the CSF-1, the mouse macrophage growth factor. J. Immunol. Methods 182: 73-79, 1995. PubMed: 7769247

Wilson CM, et al. Immortalization of growth factor-dependent mouse splenic macrophages derived from cloned progenitors. J. Immunol. Methods 137: 17-25, 1991. PubMed: 1707081