role of BSA in chemotaxis - (Apr/02/2008 )

Hi!
I am trying to do chemotaxis experiments using a boyden chamber (from neuro probe) and I have a doubt with the protocol.
I read that some authors resuspend the cells in 0,1% BSA before plating them into the chamber, but some othe authors do not. Is it neccesary to do that? Why? Which is the role of BSA?
I would also like to test the effect of some inhibitors on PDGF-BB stimulated migration. Shoud I pre-treat the cells with the inhibitor/s before plating them? Moreover, should I add the inhibitors to the upper chamber with the cells or to the lower one with the inductor, PDGF-BB?
Thaks in advance for your help.
All the best.

Mario

Hi!
I am trying to do chemotaxis experiments using a boyden chamber (from neuro probe) and I have a doubt with the protocol.
I read that some authors resuspend the cells in 0,1% BSA before plating them into the chamber, but some othe authors do not. Is it neccesary to do that? Why? Which is the role of BSA?
I would also like to test the effect of some inhibitors on PDGF-BB stimulated migration. Shoud I pre-treat the cells with the inhibitor/s before plating them? Moreover, should I add the inhibitors to the upper chamber with the cells or to the lower one with the inductor, PDGF-BB?
Thaks in advance for your help.
All the best.

Mario

I think the BSA is added to protect the cells and whatever growth factors or similar that you are using in your assay, now that you have removed all their serum. It cant harm....
As for inhibitors, it probably depends on which inhibitor you are using, how long they take to work. I think you just have to experiment a bit and see how you get the best results. As for starting conditions I would pretreat the cells for 1-2 hours before running the assay and include it the upper chamber with the cells. But be careful with inhibitors, a lot of them are also just a bit toxic generally, so be sure to include controls to show that your cells are ok, ie. will still chemotaxis normally towards another attractor.