tract tracing +immuno

Viktor Kharazia kharazia at MED.UNC.EDU
Thu Apr 18 16:50:24 EST 1996


I have a several suggestions:  (1) as a tracer use HRP(WGA-HRP) =

process with DAB, DAB/Ni,Co, TMB-tungstate or molybdate, etc., combine =

with immuno visualizing with nannogold probe: 1nm gold-anti(whatever =

animal) ( final step is silver enhancement). It is effective but =

little bit expensive. To eliminate glutar!!! from tissue before immuno =

use pretreatment with sodium borohydride.
(2) CTB-gold is an excellent tracer compatible with all kind of immuno =

(florescence or light  DAB). I am not sure if it still on a market. =

For gold enhancement use silver from Amersham and pretreatment with =

sodium metaperiodate.
High glutar making tissue almost impermeable for antibody and rising =

backgroung. To eliminate glutar!!! use pretreatment with sodium =

borohydride before immuno: 1% 30 min, then 50% alcohol 20-30min
It=B9s not clear for me why such high glutar you use. In principle, 1-2% =

is enough for everything (e.g GABA will survive even under 0.1%). Low =

glutar can be quenched with sodium borohydride, and florescence can be =

applied. =

(3) HRP(WGA) use as a tracer, process peroxidase with DAB or TMB, =

embed into resin (Epon, Durcupane, do immunoperoxidase or immunogold =

on a semithin sections (not for proteins) or do immunogold on a thin =

sections (not for every protein).



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