COMBINATION OF IMMUNOCYTOCHEMISTRY AND INSITU HYBRIDIZATION IN THE SAME SEMITHIN SECTIONS - DETECTION OF MET-ENKEPHALIN AND PROENKEPHALIN MESSENGER-RNA IN THE HYPOTHALAMIC MAGNOCELLULAR DORSAL NUCLEUS OF THE GUINEA-PIG
V. Mitchell et al., COMBINATION OF IMMUNOCYTOCHEMISTRY AND INSITU HYBRIDIZATION IN THE SAME SEMITHIN SECTIONS - DETECTION OF MET-ENKEPHALIN AND PROENKEPHALIN MESSENGER-RNA IN THE HYPOTHALAMIC MAGNOCELLULAR DORSAL NUCLEUS OF THE GUINEA-PIG, The Journal of histochemistry and cytochemistry, 40(4), 1992, pp. 581-585
We describe a procedure for combining pre-embedding peroxidase immunoc
ytochemistry with pre-embedding autoradiographic in situ hybridization
in the same vibratome sections of paraformaldehyde-fixed brain tissue
. The simultaneous detection of Met-enkephalin (Met-enk)-immunoreactiv
e product and pro-enkephalin (PE) mRNA in neurons of the magnocellular
dorsal nucleus (MDN) in the guinea pig hypothalamus was carried out a
s a model for this procedure. Vibratome slices were processed for Met-
enk immunodetection followed by the incubation with a 45-base syntheti
c oligonucleotide complementary to PE mRNA labeled with S-35. Tissues
were embedded in araldite, cut into semi-thin sections, and processed
for autoradiography. Many neurons double labeled for Met-enk and PE mR
NA were viewed in the MDN. The histological quality and the spatial re
solution of both signals were optimized, since precise intracellular l
ocalization of hybridization sites was possible. This method allows si
multaneous study of peptide immunoreactivity and mRNA expression level
s in neurons within the same semi-thin sections. It may be useful for
a variety of quantitative analyses, and might also be extended to ultr
astructural analysis.