please help with HMEC cultures - never pass 2 times... (Aug/14/2009 )

i'm culturing my HMEC in gelatin -coated flasks (by me) using MCDB medium and EGF, heparin and FBS (10%) and the cells did not attach... what should I do?

anybody knows anything special about HMECl... or any idea about problem of cell loss
thanks

I use Greiner tissue culture treated flasks for growing HMECs. The cells attach very beautifully in spite the fact that I do not use serum in the media. I use DME-F12 with growth factors recommended for HMLE cells (Human breast cancer cells generated by oncogenic transformation of primary mammary epithelial cells.Elenbaas B, Spirio L, Koerner F, Fleming MD, Zimonjic DB, Donaher JL, Popescu NC, Hahn WC, Weinberg RA.Genes Dev. 2001 Jan 1;15(1):50-65.)

i should use:

CELLCOAT®
Protein-coated culture vessels for adherent cell culture. Our CELLSTAR® product family is coated with the following proteins: Collagen Type I, Poly-D-Lysine, Poly-L-Lysine

and w' protein are your flasks coated...???