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Coating plates for improving 293 cells adherence


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#1 Tiny

Tiny

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Posted 12 November 2009 - 08:32 AM

Hello, we're in search of an adherence molecule to coat plates for HEK293 cells. We've tried Poly-lysine and it works great. Unfortunately, we also need to assay surface protein levels of these 293 cells by surface biotinylation. The biotin molecule specifically binds to an amine group on lysine and this allows non-specific labeling of all proteins on the cell surface (assuming every protein would have lysine residues). However, as you can imagine, coating the plate with poly-lysine creates this disastrous competition between the surface proteins and the poly-lysine... Therefore, an ideal coating material for us should fulfill the following criteria:

1. Improve adherence of 293 cells to an extent that is at least as good as poly-lysine (gelatin does not work very well).

2. Does not strongly compete with surface protein to be labeled with biotin.

3. Does not strongly activate signaling pathways such as integrin signal as the 293 cells will be assayed for activation of another signaling pathway that have cross-talks with integrin signaling. This sounds very hard as a lot of the coating molecules such as fibronectin and collagen would activate integrin signaling, although I'm not sure to what extent they do so. Could someone lend their expertise on this?

I also noticed that someone mentioned coating their plates with FBS in this forum. How does this work?

Any suggestions/protocols would be highly appreciated!




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