In situ hybridization (ISH) is a molecular biologic technique for revealing the location of specific DNA and RNA sequences in tissues, cells, chromosomes, or other structures by hybridizing the complementary strand of a nucleotide probe to a particular sequence. ISH and immunohistochemistry (IHC) are similar in that both use samples fixed on microscope slides and require interpretation under the microscope. However, IHC relies on protein chemistry, while ISH relies on nucleic acid chemistry for target detection.
In a hybridization assay, probes may be single- or double-stranded nucleic acid prepared from purified genomic material, recombinant fragments, or chemically synthesized oligonucleotides. Factors that must be considered in the selection of a probe for ISH include sensitivity, specificity, ease of penetration of the probe into tissue, stability of hybrids, purpose of application, and reproducibility of the method. Biotin, fluorescein, digoxigenin, alkaline phosphatase, and bromodeoxyuridine are commonly used for labeling a probe. Fluorescent in situ hybridization (FISH) is an effective technique that enables direct visualization of genetic alterations in cells. However, FISH requires a specialized fluorescence microscope and its signal fades with storage. Chromogenic in situ hybridization (CISH) is an appropriate alternative method and overcomes the disadvantages of FISH assays.
ISH is superior to analytical methods that rely on extracted samples, when cells containing the target of interest are few in the sample and histology or cytology is key to interpreting the results. ISH enables maximum use of tissue that is difficult to obtain. Hundreds of different hybridizations can be performed on the same tissue. In addition, there are two main methods to improve the sensitivity of ISH. One is to amplify target nucleic acid sequences before ISH and the other is to amplify the signal after completion of hybridization.
Amerigo Scientifics offers in situ hybridization probes, detection kits, and related reagents for localizing specific nucleic acid targets within chromosomes, cells, and fixed tissues.
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