In Situ Hybridization (ISH) is fundamentally different than Immunohistochemistry (IHC) in the sense that ISH localizes the DNA or RNA strand, whereas IHC is looking at individual proteins. With proper conditions in the laboratory, the binding process will occur spontaneously. This is a key step in DNA fingerprinting.
What is this particular method used for?
ISH offers insight into physiological processes and disease pathogenesis. It requires many steps to be taken with precise optimization for every tissue used, along with fixatives such as formaldehyde. It’s extremely useful in discovering more about the regulation and functions of genes along with the structure of chromosomes.
What is the Process?
For hybridization, cells and tissues are first treated to put the target transcripts in place and give better access to the probe, which is either a complimentary DNA or RNA. At an elevated temperature, the probe hybridizes and the excess is washed away. Once the probe was labeled, it is either localized or quantified using autoradiography, fluorescence microscopy or immunohistochemistry.
The common tissue sections that are used for in situ hybridization are:
- Frozen sections- these tissue samples are quickly frozen and once they are, they are put into a special support medium for thin cryosectioning. Then a small percentage of paraformaldehyde is used just before hybridization.
- Paraffin sections- these are fixed just like you would fix tissue for histology and then are embedded in wax before they are sectioned.
- Cells in suspension- cells can be cytospun in onto a glass slide and prepared with methanol.
With DNA Fingerprinting, being able to isolate sections of DNA will be crucial for deciphering what certain nucleotide patterns mean, and what they could mean when it comes to diagnosing diseases and preventing them.