Ultrastructure of in situ hybridization

Techniques for the ultrastructural localization of structures identified by in situ hybridization are being developed for both preembedding labeling and labeling on thin sections (postembedding). Successful labeling of both RNA and DNA sequences has been reported in recent years. Biotinylated nucleic acid probes are becoming increasing available. Colloidal gold is the only successful ultrastructural label with meaningful spatial localization, and the best results have been obtained with small (20-5 nm) gold particles. The link between biotinylated nucleic acid probes and gold has been protein A, antibiotin, or avidin binding. The size of the target nucleotide sequence, the size of the probe, and the number of gold particles attached to the labeling protein must be understood before there can be meaningful interpretation of micrographs. In addition, the spatial considerations depend on whether preembedding or postembedding is used.