Fixation for phalloidin staining - What fixative should be used for phalloidin? (Jul/15/2008 )
Hi,
We're doing an experiment to visualise actin with phalloidin rhodamine (Molecular Probes R415), and are having problems fixing with 4% paraformaldehyde; the cells have bad morphology and there is a lot of large detrius clogging the field of view.
Has anyone used methanol to fix cells for phalloidin staining? How? We've read that methanol is not a good fixative for phalloidin but can anyone tell us that they've used methanol with good results, or a reason why methanol isn't suitable?
Thanks so much!
>Shresh Pathak and Pranai Tandon
-shresh-
QUOTE (shresh @ Jul 15 2008, 03:25 PM)
Hi,
We're doing an experiment to visualise actin with phalloidin rhodamine (Molecular Probes R415), and are having problems fixing with 4% paraformaldehyde; the cells have bad morphology and there is a lot of large detrius clogging the field of view.
Has anyone used methanol to fix cells for phalloidin staining? How? We've read that methanol is not a good fixative for phalloidin but can anyone tell us that they've used methanol with good results, or a reason why methanol isn't suitable?
Thanks so much!
>Shresh Pathak and Pranai Tandon
We're doing an experiment to visualise actin with phalloidin rhodamine (Molecular Probes R415), and are having problems fixing with 4% paraformaldehyde; the cells have bad morphology and there is a lot of large detrius clogging the field of view.
Has anyone used methanol to fix cells for phalloidin staining? How? We've read that methanol is not a good fixative for phalloidin but can anyone tell us that they've used methanol with good results, or a reason why methanol isn't suitable?
Thanks so much!
>Shresh Pathak and Pranai Tandon
Methanol disrupts cytoskeleton, hence.
Phalloidin-rhodmaine works great with paraformldehyde. You need to make sure your 4%P is made fresh, and that there are no other technical toubles.
-cellcounter-