What does ISH mean in PHYSIOLOGY

In situ hybridization (ISH) is a laboratory technique used to detect and localize specific DNA or RNA sequences within tissues. The technique involves the labeling of a nucleic acid probe with a marker molecule such as fluorescent dye, biotin, or radioisotopes and then exposing the tissue to the labeled probe. By detecting the labeled molecules in the tissue, ISH allows scientists to determine which cells and cell types express particular genetic sequences.

ISH works by labeling a single-stranded DNA or RNA probe with either radiolabels, fluorescent tags, or enzymes that can be visualized under a microscope. The labeled probe is then applied to a sample of tissue containing both fixed cells and unfixed complex extracellular matrix (ECM). If the sample contains any sequence complementary to the probe, it will bind to it; when viewed through a microscope the bound probe can be detected using various visualization techniques, such as fluorescence microscopy or autoradiography.

One key advantage of ISH is its ability to detect low levels of expression; whereas previous methods tended to require highly expressed genes in order to be detectable. Additionally, unlike other DNA/RNA detection methods such as Northern blotting and reverse transcriptase polymerase chain reaction (RT-PCR), ISH gives scientists spatial information about gene expression that cannot be replicated with other techniques. Finally, ISH can detect multiple genes at once by applying several labeled probes which allows for an efficient analysis of complex cellular systems.

ISh enables scientists to study gene expression patterns at both low and high resolution levels in both live and fixed tissues. At the highest resolution level possible with microscopic imaging systems, individual cells can be monitored for precise localization and quantification of specific mRNA transcripts within subcellular regions like nucleus or cytoplasm. Furthermore, whole organisms can also be examined on how different cell populations interact with each other within their native environment via 3D imaging microscopy applications such as CLEM (correlative light electron microscopy).

In situ hybridization is typically used when one wants to observe where exactly certain genes are expressed in an organism’s body. This method generally provides higher resolution than traditional cloning/sequencing approaches because it allows gene expression patterns within individual cells and across entire organisms to be observed and quantified simultaneously with spatial accuracy significantly higher than most other molecular techniques available today.

Detectors used commonly with in situ hybridization include radioactive isotopes such as gamma emitters like 32P which provide high sensitivity but short half-life; fluorescent dyes such as Cy3 that absorb light within visible frequencies providing moderate sensitivity over long periods; and enzymes like horseradish peroxidase attached directly onto probes which provide moderate efficiency over long periods.

Samples used for this procedure must first undergo fixation via cross-linking techniques such as formaldehyde vapor treatment or overnight incubation in 4-10% buffered formalin solution prior to application of probes so that they remain intact after treatment given their labile nature due to frequent denaturation during handling steps. After fixation process has been completed then samples are often dehydrated into paraffin blocks before carrying out staining protocol.