Lipofectamine transfection, Flow sorting & cell viability??
Sze Wah Agnes CHAN
chansw3 at hkucc.hku.hk
Wed Mar 19 22:46:11 EST 2003
Dear all, greetings!
I've done Lipofectamine transfection on an established tumor cell line.
24-hrs post-transfection I've flow-sorted the GFP+ cells and plated them out.
My questions are:
1. After transfection, most of the usually attached and spread out cells
rounded up. Many became floating but most, though round-up, remained
Is this normal?
2. I've aspirated the floating cells (thinking they're likely dead) and
flow-sorted only the attached cells. Normally these cells will all attached
and spread out overnight. However, the transfected and flow-sorted ones
didn't with only countable few cells attached overnight. I did notice some
apoptotic bodies and granular "dying-looking" cells among the floating
population. However the proportion of them was small. Then I did a quick
trypan blue staining of the floating cells and saw close to none blue dead
cells. ??!! Is it usual that it may take longer for the cells to attached
after the above processes? Likely due to damages of the cell membrane? If
so, any idea how long it takes for the cells to recover?
Many thanks for advice.
Department of Pathology
The University of Hong KOng
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