Western problem: bubbles forming INSIDE PVDF membrane - (Sep/05/2007 )

I'm doing a round of Westerns and have just encountered a most unusual problem in two of my four PVDF membranes. Bubbles have appeared INSIDE the PVDF membrane. They are definitely not on the surface: they are visible from both sides of the membrane, and if I poke them with my forceps I can make them move around, join with other bubbles, etc. I am further confused by the fact that the other set of transfers I did on the same day, using the same buffers, membranes from the same lot, and transferred using identical conditions, do not have this problem.

To clarify, this is not a problem of bubbles appearing on the surface of the membrane during staining with antibodies. The membranes are currently in blocking buffer, and the bubbles seem to be completely inside the PVDF.

Has anyone else ever seen this before??? I tried searching the archives, but couldn't find anything similar.

Any insight as to what has just happened would be greatly appreciated!!!

Thanks,

Ginger

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How old is your membrane??? I have never heard of such a thing! curios! Had someone ever this problem before? DId the membranes come in contact with something which could damage the surface? Something acidic? Strange...

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Just wondering...

Are you using one sheet of PVDF membrane?

Not two sheet that sticked together??

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Nope. Definitely one. And to make things even stranger, the bubbles seem to be slowly going away again as mysteriously as they appeared. They're still there, but there aren't as many as before. I'm still a bit worried about how the blots are going to look. I'll just wait and see.

Grrr. It's just not been my week.

Ginger

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if you had been using a supported nitrocellulose then the answer would be that part of the membrane became detached from the support and allowed a bubble (which is slowly leaking out) in.

i don't know if your pvdf is constructed similarly but, if it is, then this may be what happened.

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