tract tracing +immuno

[Bo de Lange]

In article <3165CA69.109B at ere.umontreal.ca>, Denis Boire <boired at ere.umontreal.ca> wrote:

> I am looking for a retrograde tracer that could be utilized in combination with 
> immunohistochemistry which requires a fixation with 5% glutaraldehyde and revealed with 
> a peroxidaze reaction. Since so much glut is involved ...fluorecent tracers seem to be out of 
> the question. I am condidering cholera toxin B or biocityn and doing a double immuno 
> labelling. HRP labelling for neurotransmitter immuno and alkaline phosphatase labelling 
> of the tracer. 
> Has anyone experience with this combination


I've done double immunos with alkaline phosphatase and DAB visualization of peroxidase and find that if you've got a lot of deposition for both, it's hard to tell if you've got a double signal. I've successfully used Nickel-Lysine as a retrograde tracer (develops into blue-greyish) in combination with peroxidase-DAB for immuno. Best results are obtained sectioning and photographing the backfill before doing the immuno. Don't know about the 5% GA though, sounds like an awful lot to me. I used a 1-2-4% stepw
ise paraformaldehyde fix for my exps.



Bo de Lange