Brain blotting: a method to detect multiple DNA copies in specific brain regions.
We developed a method to detect multiple DNA copies (both cellular and viral) in specific brain regions by blotting thick frozen sections onto nylon membranes. This was achieved by "printing" the frozen sections on standard blotting paper immediately after cryotome sectioning and performing blotting according to the standard Southern technique. A "replica" of the blotted section was obtained by keeping on the glass slide the next frozen section cut, which was then stained for conventional histopathological analysis and the cell nuclei counted to give an estimate of the total amount of DNA present in each section. The blotted membranes were then denatured and hybridized with a nick-translated Alu probe either at 42 degrees C with 50% formamide or at 68 degrees C without formamide. Brain sections from mice infected with Herpes simplex virus 1 (HSV1), blotted and hybridized with a nick-translated HSV1 probe, clearly showed the focal nature of the Herpes simplex infection, which was also demonstrated immunohistologically using a virus specific antiserum. This method of DNA detection, conveniently modified, might also be used to detect nuclear and cytoplasmic RNAs in specific coronal sections of whole brain before localization at high power by standard in situ techniques.
Hernandez Bronchud
M
M
Department of Clinical Neurology, Radcliffe Infirmary, Oxford, United Kingdom.
Webb
S
S
Esiri
M M
MM
D000818:Animals; D001923:Brain Chemistry; D004247:DNA / Q000032:analysis*; D005629:Frozen Sections; D006801:Humans; D051379:Mice; D009693:Nucleic Acid Hybridization; D014176:Protein Biosynthesis; D051381:Rats
D004247:DNA