Immunocytochemical detection of 5'-bromodeoxyuridine in fluoro-gold-labeled neurons: a simple technique to combine retrograde axonal tracing and neurogenetic characterization of neurons.
To characterize the axonal projections of 5'-bromodeoxyuridine (BrdU)-labeled neurons, we have combined retrograde tracer injection of Fluoro-Gold with the immunocytochemical detection of BrdU. Pregnant mice were labeled with pulses of BrdU at embryonic days E12, E13, E14, or E16. Young adult offspring were perfused with 4% paraformaldehyde 2 days after receiving a Fluoro-Gold injection into the cerebral cortex, thalamus, or hippocampus. Brain sections were processed for immunocytochemical visualization of BrdU using the peroxidase-anti-peroxidase method and a diaminobenzidine-nickel ammonium sulfate (DAB-Ni) reaction, and finally observed on a microscope equipped with brightfield and fluorescence optics. Both BrdU-immunoreactive nuclei and retrogradely labeled Fluoro-Gold-positive cells were detected. Double-labeled neurons were recognized by the presence of fluorescent particles in the cytoplasm and a black immunoreactive nucleus. Since both labelings occurred in different cell compartments, Fluoro-Gold granules were not obscured by the DAB-Ni precipitate. The method shown here permits a correlation of the neurogenesis of subsets of neurons identified by their BrdU content with the specific target into which such cells project.
Unidad de Biologia Celular, Facultad de Biología, Universidad de Barcelona, Spain.
E
E
Soriano
J A
JA
Del Río
F J
FJ
Martínez-Guijarro
I
I
Ferrer
C
C
López
D000818:Animals; D001369:Axons; D001973:Bromodeoxyuridine / Q000378:metabolism*; D005260:Female; D007150:Immunohistochemistry; D051379:Mice; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism*; D011247:Pregnancy
D001973:Bromodeoxyuridine