Fluorescence in situ hybridization (FISH)

Fluorescence in situ hybridization (FISH) is the methodology used to detect specific nucleotide sequences by using DNA probes labeled with a fluorochrome that will hybridize with denatured chromatin (DNA) on a microscope slide. The process involves the breaking of hydrogen bonds during DNA denaturation or dissociation, and then reannealing the broken strands, during hybridization, in the presence of labeled DNA single strands of the same sequence. This simplified process allows the targeted genetic sequences to be visually examined under a fluorescent microscope that is properly equipped for that fluorochrome. Because this technology can be successfully used on non-dividing nuclei to detect aneuploidy or rearrangement, it is not restricted to mitotic cells, and thus can be used on paraffin-embedded breast tissue or free-floating amniotic cells. And because it does not require culture time to obtain dividing cells, it also eliminates any culture delays that might otherwise be required if chromosomes were required. The authors of Chapter 16, Fluorescence In Situ Hybridization, in the AGT Cytogenetics Laboratory Manual, 4th ed., provide not just a comprehensive review of the theoretical mechanism that is occurring at the submicroscopic DNA level, but also offer a wealth of history and troubleshooting tips that reflect their years of hands-on experience in this area. Contributed protocols are included with this chapter.