Anyone can give me some advice in culture THP1 cell line?
I have a trouble in thawing THP-1. Why they almost died ?
I thaw a vial of THP-1 cell line to the 10 cm dish. The vial contain 10^7cells in 5%DMSO.
The procedure of thawing cell, quickly thaw in water bath,then mixed with 3 ml fresh medium, and centrifuged with 300g 3 mins, to remove DMSO.
Even they almost died, but still a small protion alive. The reason I know they still have little portion alive is, I placed them in RPMI1640 more then 2 weeks, they gradually form many colonies suspension in medium.
So the next question : Does these THP-1 cell are normal? Does these cell suitable to different M0 M1 and M2 macrophage?
(I suppose after we treat with PMA, the cell is in the M0 state.)
Thank you
Dear Ting-Wei Hsu
please see the attached link for details concerning the propagation and differentiation of THP-1 cells. I have used the Bowdish lab protocols with great success and can only recommend them.
http://www.bowdish.ca/lab/protocols/macrophage-isolation-culture/
All the best & kind regards,
Michael
We have also struggled with this in the past.
We found difficulty thawing if the cells were not frozen down in high FBS concentrations (90%).
When thawing in the water bath i always make sure they are never fully thawed. Leave a small remaining frozen section until you transfer the cells.
I personally used 10mL media at 150g x 10min to remove DMSO.
These cells are fragile, they will have a relatively high death rate and they grow slowly. I would often culture mine for almost a month before I would use them.
Might be problem with frozen media. You can try to add media and put cells into culture small culture flask for some time (1 day or 6-8 hrs) and then you can spin down at low rpm because cell is very fragile and transfer into fresh media but keep media in small volume otherwise it will give shock to the cells. Hope it will work.
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