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[Suzanna]

I think I must be really stupid but my effeciencies with calcium phosphate actually work better than fugene/lipofectamine!! What is a typical protocol for fugene transfection ? does adding fugene to the diluent with dna make a difference than making separate dilutions of dna and fugene in serum free medium and mixing ? also what is the typical complexing time ? what efficiency should i expect for an endothelial cell line (p9)? any tips and suggestions welcome

thanks

Suzanna

[bob1]

Fugene in the presence of water forms small charged droplets called lipospheres with the DNA inside them due to charges on the DNA that are not present in the medium/water (medium is mostly water).  You should be adding the fugene directly to the DNA/diluent mix.

Complexing should be 15-45 min, but can be longer.

Can't comment on the endothelial line though.