I want to check the TLR4-mediated activation of NF-kB in response to LPS. So, I transfected my DOV13 cells with CCL20 for 24h. I did this in 24-well plate, however I had problem with the cells, the last half from the 24 well-plate were detached after 24h transfection, but confluence or cell density is not a problem as there is still space on the plate. So, I worked on the remaining halves which has good attachment, but I detected very low data.
Now, I really cannot tell which procedure I did wrong, by the way, I used Lipofectamine 2000 for transfection and I used the lowest volume possible (0.5 ul/well). Thanks.
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