Dear Mohamadreza
Please see this procedure
http://www.ncbi.nlm.nih.gov/pubmed/1290726
Dear Mohamadreza,
I haven't freeze macrophages directly, but I had several time freeze bone-marrow to then, later on defreeze it when required and differentiate into bone marrow derived macrophages. For sure the final amount of macrophages you have is fewer but still, it is worth for it.
Briefly, once you flushed the bone marrow, centrifuge it (5' 1500rpm) and resuspend it in FBS with 10% DMSO to then freeze it in criovials. Per mouse I usually generate two vials.
Hope this works for you.
Hi Mohammadreza,
I have worked with alveolar macrophages extracted from lung and when I freeze them I usually use the next freezing medium: 40% complete culture medium, 40% FBS and 20% DMSO. I mix an equal volume of cell suspension with it, aliquot and freeze.
hi gabriela
and do you measure or control their activity and number after defrost?
After thawing, I usually check viability and have ~90-95% live cells and in terms of activity they give nice responses to TLR agonists ( LPS or PolyIC)
do you know that remain active without extracting or not?
i.e in lung or liver freezing
hi shahida
you have never frozen macrophages means its not possible macrophages be freeze?!
There are number of papers where they tried freezing macrophage. Pleae check the following paper.
Freezing of rat macrophages; E. Hem, A.C. Munthe-Kaas
look here
https://lists.purdue.edu/pipermail/cytometry/2012-May/043481.html
Hi,
Want to grow primary rat Kupffer cells culture (liver macrophages) in collagen coated 24 well plate. We have cryo-preserved cells. Could see good number of cells after seeding them in wells at density 0.4 million/well. After 24 hrs, I can see loose aggregates and few attached cells (with round/granular morphology). After replacing media, only few cells are observed in well. What could be the reasons?
Thanks in anticipation
Well, someone is able to do it. PromoCell actually sells frozen human macrophage differentiated with CSF. They do not describe their freezing method.
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