[Histonet] Problems with Gold-immunolabeling and silver enhancement in brain tissue

[Adam Perry]

I'm trying to perform silver enhancement of my gold
labeled secondary antibody (1.4nm gold) in an
immunocytochemistry (ICC) protocol in rodent brain
tissue.  I know that the primary antibody and initial
steps are working because when I use an
avidin-biotinylated peroxidase kit to visualize a
biotinylated secondary antibody (as opposed to using
the gold conjugated secondary antibody)I get great
labeling of my antigen.  I've done a dot blot with the
gold labeled antibody and I seem to get silver
enhancement of particles only in the dot blot with the
gold-labeled antibody (as opposed to unlabeled
antibodies)- so I'm pretty sure there are gold
particles attached to the IgG.

Are there any obvious differences that need to be
considered when working with a gold-conjugated
antibody as opposed to antibodies that are
biotinylated or conjugated to enzymes??      

Any insights would be greatly appreciated..
Thanks,
Adam Perry
Department of Physiology and Biophysics
University of Illinois
Chicago, IL

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