I have a couple of questions regarding primary NK cell isolation from human peripheral blood:
What is the practical yield of NK cell isolation from human blood?
(for instance using the Miltenyi Biotec MACS NK cell isolation kit)
Is it possible to expand these cells in culture? Does anyone have any protocols? How long will primary NK cells stay alive in culture?
Is there any cell line of human NK cells for NK activation, cytotoxicity and functional studies?
Many thanks in advance!
Brian summed it up nicely.
We also added 3.5µl beta-mercaptoethanol per ml culture medium.
We cultured them for a max of 2 weeks.
If you want you can also select for adhering NK-cells you can use the following culture medium:
RPMI
10% hAB serum
1 mM CaCl2
1mM MgCl2
100 u/ml heparin
IL-2 as you are used to.
After 24 hours culture was aways non-adherent cells and continue culturing using same medium.
From PBMC, I think you can get up to 15% of NK cell population. In our lab, we isolate NK cells from PBMC using STEM CELL Technologies negative selection of NK cells and afterwards, we do the same as what Brian mentioned: we culture in RPMI 20% FBS plus IL-2 and IL-15 and culture them for 2 weeks.
We also use NK-92 in the lab, and followed ATCC's recommended medium https://www.atcc.org/products/all/CRL-2407.aspx#culturemethod plus IL-2 and IL-15.
The MACS isolation kit works fine. 95% purity of NK-cells was reached easily.
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