Does high osmolarity help antibodies bind epitope?
Immunodevice at yahoo.com
Sun Oct 13 15:13:23 EST 2002
I would imagine they were trying to achieve a salting-out effect, to get
better apparent binding - q.v. high salt buffers for affinity purification
of Ig's on Protein A. This may also increase non-specific binding. Why not
try with and without NaCl, then use the "best"!
<tmorris at uhnres.utoronto.ca> wrote in message
news:20021008180348.17091.qmail at ww02.hostica.com...
> Are there any experts on the practical aspects of immunocyt? If so I thank
you in advance for advice on this one.
> Our lab has a hand-me-down recipe for antibody buffer, for staining
imobilized isolated neurones for fluorescent microscopy. It's basically a
tris based buffer which includes 500mM NaCl. No one can offer a rational
explanation for this, nor can I find anything similar on the web or in
books. I've used a borate buffer with a more normal osmolarity (300 mOsmol)
and the staining was just as good. I suspect PBS would work also, which is
what most books reccomend.
> Such high salt might neutralize static charges on IgG, but why would this
help in binding epitope. Surely, the closer to physiological conditions the
better. Am I missing somthing? The person who designed this buffer must have
had a good reason and I don't want to just ditch a protocol without a clear
understanding of the whys and wherefores.
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