Hi Philip,

in our lab inorder to avoid fibroblast contamination we use to preplate culture solution for 1-2 our in normal plastic, (fibroblasts quickly adhere), and then harvets the supernatant for target cells plating.

Good Luck

Try to eliminate fibroblasts by panning onto a Thy-1 coated substrate, immediatly after having prepared your suspension.

Great! thanks guys

Phillip, I hve tried 2 methods to control fibroblast with success. Plate the endothelial/muscle cells with fibroblast for 1 hr. Then aspirate the media into a fresh cell culture flasks. Fibroblast will adhere to the plastic in the first cell culture flask, you may need to this more than once.  A second method after aspirating the media from the original flask after 1 hr, decrease the concentration of the FBS to 1-5%. Endothelial cells will proliferate at this concentration of  FBS. I suspect muscle cells will also proliferate at 5% FBS.

A third method which will deliver cleaner cultures is remove the media, add PBS to remove FBS proteins, aspirate PBS, add 1% solution of EDTA:Versene for 3 minutes. Fibroblast will lift off plastic, endothelial cellsremain attach. Aspirate EDTA:Versene, rinse with PBS, then add media with low FBS concentration. Essential point is limiting the EDTA:Versene to 3 minutes.

I would mean in my first comments "onto an anti-Thy1 antibody- coated substrate". We use with success this méthode to eliminate fibroblasts from dissociated sciatic nerve, in order to purify Schwann cells.