How to prepare conditioned media? - Do I have to omit supplements for the culture media? (Dec/05/2007 )

Hi all,

I need to prepare conditioned media, add this to other target cells and see whether this conditioned media cause apoptosis of target cells.

I use RPMI or Keratinocyte-serum free media but I am not sure whether I have to add serum or supplements to the media (EGF and bovine pituitary extract are two

supplements for Keratinocyte-SF media). I just guess growth factors (serum/supplements) may protect target cells from apoptosis, so I want to know how usually

people prepare conditioned media (with or without serum/supplements???? or it depends on the purpose of the experiments???).

Thank you so much in advance!!

I haev done similar expriements and and added conditioned media plus a
small volume of non-conditioned. For example to 12 well plates I added 1.5 ml conditioned media + 1 ml non-conditioned to help keep cells alive.
However, this will have to be determined empirically. It is possible that others may have done something similar in the literature.

I think this depends on the purpose of the experiments. What do you wanna show ? Maybe experiments with conditioned media are not the suitable experiments.... you should at least try both,as control.