need help on retrograde labeling

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traceruser
traceruser's picture
need help on retrograde labeling

Hi All,

I am a graduate student in Boston University. The project I am working on includes retrogradely label mitral cells in the mouse main olfactory bulb. I use CTb and it work well (tracer injection injected to amygdala). I also want to distal dendrite being labeled. Because the dendrite is very long and the end of the dendrite is not well labeled. I wonder whether there is better way to back label distal dendrite.

Thank you very much !

Cam ( eval(unescape('%64%6f%63%75%6d%65%6e%74%2e%77%72%69%74%65%28%27%3c%61%20%68%72%65%66%3d%22%6d%61%69%6c%74%6f%3a%63%61%6d%6d%79%40%62%75%2e%65%64%75%22%3e%63%61%6d%6d%79%40%62%75%2e%65%64%75%3c%2f%61%3e%27%29%3b')))

Fraser Moss
Fraser Moss's picture
does this reference help?

does this reference help?

1: J Neurosci Methods. 1995 Mar;57(1):81-91.

Neuron labeling by extracellular delivery of horseradish peroxidase in vivo: a
method for studying the local circuitry of projection and interneurons at
physiologically characterized sites.

Benson TE, Voigt HF.

Department of Biomedical Engineering, Boston University, MA 02215-2407, USA.

An anatomical method is described that yields individual neurons with
continuously labeled dendrites and axons following the extracellular deposition
of horseradish peroxidase (HRP) at neurophysiological recording sites in vivo.
The method is a logical evolution of previous methods for iontophoretic delivery
of HRP: Parameters critical to the ultimate concentration of HRP at the labeling
site are reduced by an order of magnitude relative to standard practice. In
successful cases one neuron or two in the immediate vicinity (50 microns) of
recording sites is/are labeled. Labeling of other processes traversing the
injection site, if any, is subliminal at highest light microscopic magnification.
Due to the labeling of so few cells and the absence of other labeled processes,
dendritic trees and local axonal arbors can be reconstructed without ambiguity.
In addition to recovering neurons at sites characterized with physiological
(e.g., sensory) stimuli, the method offers the further advantage of being fully
compatible with subsequent electron microscopy. Both large (> 20 microns) and
small (approximately 8 microns) neuron types and glia have been labeled.

PMID: 7791368

Fraser Moss
Fraser Moss's picture
I'm not sure if this video

I'm not sure if this video will help from www.Jove.com. Click the title to watch

Retrograde Labeling of Retinal Ganglion Cells by Application of Fluoro-Gold on the Surface of Superior Colliculus

Kin Chiu, Wui-Man Lau, Sze-chun Yeung, Raymond Chuen-Chung Chang, Kwok-Fai So
University of Hong Kong

ecolechio
ecolechio's picture
The most reliable retrograde

The most reliable retrograde tracer I've used is Fluoro-Gold (iontophoretically injected). Fast Blue/True Blue has also worked well for me in the past, but I am not sure about how it works for the distal dendrites. If you need to be able to see the cytoarchitecture in your sections with FG (eg, using cytochrome oxidase), there is also an antibody to FG that will allow you to see the labeled neurons in the processed sections.

QIN
QIN's picture
I have been using Fluorogold

I have been using Fluorogold (FG) to label retinal ganglion cells via intratectum injection into the superior colliculum. I don't think the FG could label the dendrites very nicely. FG is pretty good in labelling the soma, but not the dendrites.

I was told that DiI may be a good way to label the dendrites. You could check the pubmed to get further reference.

Good luck!