Location of Gene Expression in Tissue Sections by Hybridization Histochemistry Using Oligodeoxyribonucleotide Probes

This chapter explores synthesis, labeling, and use of oligodeoxynucleotides in the technique of hybridization histochemistry. The technique of hybridization histochemistry is simple and convenient and has been used for several years for a variety of research topics and myriads of specimens with consistent success. Oligodeoxyribonucleotides are routinely synthesized by the solid-phase method on an Applied Biosystems Inc. 380A DNA synthesizer, using phosphoramidite chemistry. The most convenient method for the routine purification of large numbers of oligonucleotides is polyacrylamide gel electrophoresis (PAGE). The choice of radioisotope used for incorporation into the oligonucleotide depends on the experimental requirements. The temperature for hybridization of sections with oligodeoxynucleotides is related to the probe length, specificity of the probe for the target mRNA, and the formamide and salt concentration of the hybridization buffer. The technique of hybridization histochemistry may be used in conjunction with immunohistochemistry for the investigation of transcriptional regulation of neuroendocrine peptides and identification of the peptide product.