I requires Natural Killer (NK) cells to be isolated from whole blood. I have attempted a few different methods but can't really decide which is better. The first method was layering the blood on ficoll hypaque and extracting the buffy layer. After this I added a specific antibody for a NK cell surface marker. This sample was then run through the flow cytometer and a sort was performed on the positive population.
The second method I tried was to simply lyse the RBC's, add the antibody and then run the sample.
I would like to ask if anybody has done any of these procedures and if it worked well for them. I think the first procedure is probably best but it requires 10x more time which I'm afriad might start being worse for the cells than the lysing solution?
Is it possible to get viable NK cells out of whole blood with the use of a lysing solution? has anybody successfully done this?
Full Version: Isolating Natural Killer cells from whole blood
I did the similar experiment before,
the problem is whichever methods you use, you will only get very small number of NK cell population at the end.
(unless you have huge amount of whole blood from the begining.)
if you still want to do so, I think it is better to do ficol, since it remove a lot of unwanted cell (and save some colume of antibody).
If you just need NK cell culture, one alternative is to use cell line NK-92 from ATCC.
It is not normal NK cells (Nk leukemia cell line), and little tricky to culture them. but I thought useful.
the problem is whichever methods you use, you will only get very small number of NK cell population at the end.
(unless you have huge amount of whole blood from the begining.)
if you still want to do so, I think it is better to do ficol, since it remove a lot of unwanted cell (and save some colume of antibody).
If you just need NK cell culture, one alternative is to use cell line NK-92 from ATCC.
It is not normal NK cells (Nk leukemia cell line), and little tricky to culture them. but I thought useful.
QUOTE (Stephan @ Jun 17 2009, 07:09 PM) 
I requires Natural Killer (NK) cells to be isolated from whole blood. I have attempted a few different methods but can't really decide which is better. The first method was layering the blood on ficoll hypaque and extracting the buffy layer. After this I added a specific antibody for a NK cell surface marker. This sample was then run through the flow cytometer and a sort was performed on the positive population.
The second method I tried was to simply lyse the RBC's, add the antibody and then run the sample.
I would like to ask if anybody has done any of these procedures and if it worked well for them. I think the first procedure is probably best but it requires 10x more time which I'm afriad might start being worse for the cells than the lysing solution?
Is it possible to get viable NK cells out of whole blood with the use of a lysing solution? has anybody successfully done this?
The second method I tried was to simply lyse the RBC's, add the antibody and then run the sample.
I would like to ask if anybody has done any of these procedures and if it worked well for them. I think the first procedure is probably best but it requires 10x more time which I'm afriad might start being worse for the cells than the lysing solution?
Is it possible to get viable NK cells out of whole blood with the use of a lysing solution? has anybody successfully done this?
Have you tried the Dynal NK isolation kit from Invitrogen? This uses magnetic beads and anti- CD3, CD14, CD36,
CDw123, HLA class II DR/DP and CD235a (Glycophorin A) to deplete non-NK cells. You can then stain the enriched population for NK cells and sort.
The method is quick and gives quite a clean population.
Hope this helps.
This is a "lo-fi" version of our main content. To view the full version with more information, formatting and images, please click here.