On Fri, 19 May 1995 16:57:18 EST5ED, Riitta Miettinen wrote:
 
>Hi everyone,
>We are immunostaining for
>macrophages and dendritic cells.
>and have encountered what appears to be endogenous biotin as avidin
>conjugates alone seemed to stain tissue.  Has anyone encountered this
>problem before and are they aware of
>any techniques that block endogenous biotin in tissue prior to staining?
>
>
>There is commercially available a blocking kit for endogenous biotin:
>Avidin/biotin Blocking Kit, Vector Laboratories, SP-2001.
>In some cases it helps.
>You may also try streptavidin, instead of avidin, since they say that
>streptavidin has lower affinity for biotin in eukaryotic cells.
 
 
We also have had this problem in various tissues including skin and
autonomic ganglia. In our hands the blocking kit from Vector Labs made
little difference. Streptavidin IS a bit better than avidin in most
tissues.  I also would like to know either what the endogenous ligands may
be (I doubt it is all biotin) or how you can get rid of the binding...It is
sufficiently bad in our ganglia that we cannot use biotinylated secodary
antibodies to localise synaptic boutons, especially if Cy3 is used as the
fluorophore...
Ian Gibbins
Department of Anatomy and Histology
Flinders University of South Australia
Phone:  +61-8-2045271
FAX:    +61-8-2770085