t******n
发帖数: 33 | 1 First, thank you very much for any suggestions on this problem.
I am running ADCC assay with human NK cells activated by IL-2 overnight. I
got ~100% killing of the target cells when cultured with activated NK cells
even without antibody, please refer to the data below.
________________________________________________________________
Target cells were labeled with Europium.
Target cells+ Effector cells (no antibody treatment)
average 338662
Target cells+ Effector cells + 1% Triton X-100 (Max kill)
average 321999
Target cells only
average 131699
Target cells +1% Triton X-100
average 289936
_______________________________________________________________________
I didn't have this problem using freshly isolated NK cells (use the cells
right after isolation, no overnight culture).
I really appropriate any suggestions on this issue. Thank you! | l******a
发帖数: 3339 | 2 你用了多少effector cells，1/10吗？
cells
【在 t******n 的大作中提到】
: First, thank you very much for any suggestions on this problem.
: I am running ADCC assay with human NK cells activated by IL-2 overnight. I
: got ~100% killing of the target cells when cultured with activated NK cells
: even without antibody, please refer to the data below.
: ________________________________________________________________
: Target cells were labeled with Europium.
: Target cells+ Effector cells (no antibody treatment)
: average 338662
:
: Target cells+ Effector cells + 1% Triton X-100 (Max kill)
| t******n
发帖数: 33 | 3 Yes, I used E:T=10:1. NK cells=100,000cells/well, target cells=10,000cells/
well in 96-well round bottom plates.
After overnight culture in RPMI1640+GlutaMax+Sodium Pyruvate+10%FCS+10ng/ml
IL-2 at 1.0x10e6cells/ml, I mixed the NK cells with target cells and co-
cultured for 2 hours. I got ~100% killing of target cells even without any
antibodies.
When I removed IL-2 from the medium and cultured NK cells overnight, I got
the same problem:~100% killing after co-culture with target cells for 2
hours.
But when I used NK cells on the same day (no overnight culture), I didn't
have any problem.
Thank you very much for your suggestions.
【在 l******a 的大作中提到】
: 你用了多少effector cells，1/10吗？
:
: cells
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