colin at fungus.biochem.ualberta.ca
Mon Sep 2 17:08:46 EST 1996
> Dear All:
> I use zymolyase to break down yeast cell wall and then do immunostaining.
> For some reason the zymolyase reaction only gave 10-20% dark cells under microscope.
> The conditions are following:
> 10^7 cells of overnight culture (OD596=1)
> spin to remove media
> wash 2x with 0.1M KPO4(pH6.5)/1.2M sorbitol
> suspend cells in 0.5mL 0.1M KPO4(pH6.5)/1.2M sorbitol
> add 5ul mecaptoethanol, 5ul 10mg.mL zymolyase in 0.1M KPO4(pH6.5)/1.2M sorbitol
> 30'/37degree shaking
> Can somebody offer some advices/protocols?
Try more zymolase for the same length of time you are currently using.
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