Hi there,

I'm trying to build a killing curve for HEK-293T towards hygromycin B for the following transfection experiment. I plated the cell at 1.3X10^5 cell/well in the 24 well plates the night before I add the antibiotic. The cell never reached higher than 80% confluence due to the abx until I saw the cell detachment. After 4 days of culture with hygromycin B, the wells that abx concentration was higher than 200ug/ml have cell detached and the cells were floating like a mat in the well. I took the supernatant with the cells and count them with trypan blue but most detached cells were alive. I am a bit confused if I should consider this condition as the cells were killed or they are alive? How do I set the optimal selection abx concentration if this floating cell occurs? Thank you so much for your time!

Best,

Yi