Hi, I need to seperate type of blood cells for cell culture after I want to look some protein in this cells. For example Im goinog to take only natural killer cell from blood, after I culture them, after all Im going to check the protein and give same protein and look what happen in when I mix lots of that protein with natural killer cells. However not just natural killer, B, T also gronolocytes.
Do you have any idea how can I get them. I wonder can I sorting them by flow cytometre and what is percentege of purity?
Or should I trasform them from blood stem cell or myoloid or leymhoid stem cell and can I?
wich one is more efficient and sensible?
Thank you
Hi Kadir, Yes you should cell sort them all the incidences of these cells are relatively high, the purity should be about 98%+
Regards
Gary
I think the first one is more easy and reliable.....flow cytometry...
Dear Kadir
If you want to purify natural killer cells, you must do certain important steps
1) By using 'lymphoprep' (or some other commercially available kit), purify 'mononuclear' cell fraction which will contain lymphocytes and natural killer cells. You may need to use a large volume of blood to obtain sufficient number of NK cells.
2) You will need to decide whether you want to use a positive selection or a negative selection technique or both. In order for you to employ a positive selection method by either magnetic beads or flow cell sorter, you will need to use an antibody that is unique to NK cells. That's where you are going to have problem. In a negative selection, you deplete other types of cells. Natural cells are CD3- CD4- CD8- CD19- CD56+. So you could remove B cells with anti-CD19 tagged magnetic beads and T cells with anti-CD3 tagged beads. Then use anti-CD56 tagged beads to concentrate CD19- CD3- CD56+ cells that will contain NK cells. I know it is going to be tedious.
Good luck
Siva
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