If I flush the mouse bone marrow and culture it overnight in DMEM in tissue-culture treated plates, some cells adhere and remain so after the washings. When I Accumax them they come off very easily though. Would it be correct to say that these are mostly macrophages? Can I culture primary macrophages in this way without adding any differentiating cytokines (no M-CSF)?
Most of these will be marrow stromal cells, aka "mesenchymal stem cells" (MSC). Maybe some smooth muscle and adipocytes. Differentiated adhesive macrophages might be there too, but they would be rare. In a healthy mouse, most BM macrophage potential would exist in the form of non-adhesive "monocytes" (macrophage precursors).
Usually to get a hold of adherent macrophages, the best way would be to isolate monocytes, using a technique like MACS, and then differentiate them into adherent macrophages with cytokines.
If you want to do it super-cheap, you can try to exploit the change in adhesion that's considered somewhat unique to monocyte/macrophages: First, keep the non-adherent fraction containing among other things monocytes (getting rid of MSC). Then differentiate. Then keep the adherent fraction (macrophages).
You could also induce macrophage recruitment into the peritoneal cavity using Thioglycollate and then do an overnight adherence purification. No cytokines required.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2834554/
I need specifically macrophages from BM, and primary ones, not the differentiated BMDMs.
When I FACS analyze adhered cells they are CD45+, F4/80+ and mostly Gr1- (there's small population of Gr1+) . I assume these should be either macrophages or mature monocytes. Are there any additional markers to separate monocytes from macrophages?
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology