gotta get my $%#&* cells to STICK!!

[john.hines at yale.edu]

In article <3C7C25B3.446FFD05 at gpu.srv.ualberta.ca>,
wgallin at gpu.srv.ualberta.ca wrote:

> John,
>         One possibility is that the matrix adhesion is exquisitely calcium
> sensitive (those darn integrins).  Most PBS formulations do not have
> calcium and magnesium added, and the phosphate tends to complex with any
> trace amounts. What I do if this is looking to be an issue is to add
CaCl2 and MgCl2
> to 0.5 mM final concentration in an aliquot of PBS that is being used
> for washing.  

Excellent idea for the PBS rinses.  However, even if I just change the
growth medium I lose a lot of the tubules -- they just seem to give up. 
Maybe I need to augment the Ca+2 and Mg+2 concentrations a bit in the
growth medium.


>         The other thing I have found with some cell lines (No information
> specifically on myotubes) is that at low temperature they release
> quickly from the substrate, so for those cell lines that are sensitive I
> use prewarmed solutions when rinsing, at least until they have been fixed.

That would also be a worthwhile thing to do.  I got some non-gelatin
adhesion matrices today -- I plan on trying them out this time.

Thank you for the suggestions, Warren,

John