One of the most popular questions we get here is which medium formulation is best for culturing human and mouse macrophages. We’ve tested many different media and methods over the years and have some insights to pass along.

The Effect of Culture Medium on Human Macrophages

Human monocytes are typically considered macrophages following a 5-day culture with human recombinant macrophage colony stimulating factor (M-CSF). Monocytes will not proliferate much, if at all, but M-CSF is still required for optimal maintenance in culture.

DME/F12 vs. IMDM

We ran an experiment to compare macrophages cultured in DME/F12 (a 50:50 mixture of Dulbecco’s Minimal Eagle’s medium and Ham’s F12 medium) to Iscove’s Modified Dulbecco Medium (IMDM). Here are the highlights:

  • We supplemented both formulations with 10 ng/mL M-CSF and 10% human serum.
  • After 5 days, we added LPS to one-third of the cultures to stimulate TLR4 and polyinosine:cytosine to the other third to stimulate TLR2. The remaining third served as a  negative control.
  • We collected the culture media after 48 hours and analyzed the cytokines.
  • Findings: The cells grown in DME/F12 produced much more IL-6 and TNFα than the same cells grown in IMDM. 


Human monocytes cultured in DME/F12 (left) and IMDM (right).

The Effect of Culture Medium on Mouse Macrophages

The process of culturing mouse macrophages differs slightly from the process for human cells described above.

Since mouse macrophages are cultured from mouse bone marrow cells, they develop from progenitor cells and thus proliferate, unlike human monocytes. For our mouse macrophage experiment, we prepared macrophage cultures in 4 different media: Roswell Park Memorial Institute 1640 (RPMI 1640), RPMI 1640 supplemented with hypoxanthine and thymidine (HT), DME/F12, and IMDM. Here are the highlights:

  • All 4 media contained 10% fetal calf serum.
  • We added M-CSF at a range of concentrations.
  • After 7 days, we measured cell growth using CellTiter GloTM.
  • Findings: IMDM was by far the best medium of the 4 we tested. Growth in IMDM was superior at lower concentrations of M-CSF.

When It Comes to Media, Choose Carefully

We found DME/F12 to be the best culture medium for human macrophages and IMDM to be the best choice for mouse macrophages.

Your choice of medium can affect both cell growth and function, so don’t make any assumptions. It’s best to test several different media in your own lab setting to see how each influences your experimental design.
Happy testing!

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1 Comment

  1. Thanks for the informative article about macrophages. It’s interesting to learn that that DME/F12 is a good culture medium for human macrophages. It could be interesting to learn how other types of culture medias can be compatible with other animals as well, like the mouse you mentioned.

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