Trying to optimise the Annexin v -FIT-C kit for flow cytometry. 24 hours and especially 48 hours after drug treatment i'm getting very low viability with alot of dead floating debris but a relatively weak FIT-C signal. should I be collecting dead floating cells as well when I harvest to increase my annexin signal??
Hi,
In my opinion yes as their were alive at some point.
They might be floating as a consequence of your treatment so should be included in your analysis as well.
Alexis
Yes. If you do not collect them then you will likely miss a portion of dead/dying cells. Examining only adherent cells could select for those that are healthier, thus not yet undergoing apoptosis (or resistant to it).
Hi Rick,
ALWAYS!!! I remove the medium, add trypsin the cells then mix the trypsin back into the original medium and spin them.
Kind regards,
Steve
Hi Rick,
I agree, you will miss a part of your information if you discard the floating cells. And your control condition will help you to determine what part of cell death is due to your treatment and what part is due to the normal culture cell death.
Good luck in your experiments,
Daniel
It is important you do collect the floating dead cell as they will add information to the effect of your drug treatment.
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