Recent developments in the use of synthetic oligonucleotides for in situ hybridization histochemistry.
Synthetic oligonucleotides have been used with increasing frequency as probes for the detection and study of the regulation of specific mRNAs by in situ hybridization histochemistry. These probes can be easily obtained and used by the nonmolecular biologist, and they have been shown to be effective for the study of a wide range of mRNAs in neuronal and neuroendocrine tissues. Considerations in oligonucleotide probe design, synthesis, purification, and labeling are described in this article, and current procedures for tissue preparation and hybridization are discussed. In addition, control procedures and methods for the quantitation of in situ hybridization by image analysis are discussed. Finally, the combination of this technique with immunocytochemistry and retrograde tract-tracing is reviewed. The coupling of quantitative in situ hybridization with other neuronal markers, e.g., of connectivity, provides an increasingly valuable technology for exploring the regulation of gene expression in a rich anatomical context.
Lewis
M E
ME
Cephalon, Inc., West Chester, Pennsylvania 19380.
Krause
R G
RG
Roberts-Lewis
J M
JM
D000818:Animals; D001345:Autoradiography / Q000379:methods; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D004745:Enkephalins / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis*; D010761:Phosphorus Radioisotopes; D011498:Protein Precursors / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics*; D051381:Rats; D013462:Sulfur Radioisotopes
D004745:Enkephalins; D009838:Oligodeoxyribonucleotides; D010761:Phosphorus Radioisotopes; D011498:Protein Precursors; D012333:RNA, Messenger; D013462:Sulfur Radioisotopes; C029992:proenkephalin