In situ hybridization (ISH) is a technique for localization and
detection of a specific DNA or RNA sequence in whole embryos or tissues/tissue
sections/cells using a labeled complementary DNA or RNA strand. DNA and RNA
sequences can be visualized after hybridization with labelled probes that are
complementary to the sequence of interest.
The underlying basis of In situ hybridization (ISH) is that nucleic acids, if preserved
adequately within a histologic specimen, can be detected through the
application of a complementary strand of nucleic acid to which a reporter
molecule is attached. Visualization of the reporter molecule allows localizing
DNA or RNA sequences in a heterogeneous cell population including tissue
samples and environmental samples. The technique is particularly useful in
neuroscience.
The current applications of this technique include: in situ
hybridization to mRNA with oligonucleotide and RNA probes; analysis with light
and electron microscopes; whole mount in situ hybridization; double detection
of RNAs and RNA plus protein; and fluorescent in situ hybridization to detect
chromosomal sequences.
DNA ISH and RNA ISH
DNA ISH can be used to determine the structure of chromosomes. Fluorescent DNA ISH (FISH) can, for
example, be used in medical diagnostics to assess chromosomal integrity. RNA
ISH (RNA in situ hybridization) is used to measure and localize RNAs including mRNAs,
lncRNAs, and miRNAs, within tissue sections, cells, whole mounts, and
circulating tumor cells (CTCs).
Custom ISH Service
Creative Bioarray offers completely customized ISH service from probe design and tissue procurement to
expertly interpreted gene expression studies. Its proprietary ISH techniques can
dramatically reduce operating costs by eliminating the complicated
time-consuming processes of ISH away from laboratory.
When carrying out this technique, cells and tissue sections are
typically fixed in 4% paraformaldehyde to preserve morphology for ISH. In some
cases, tissues are permeabilized with proteinase K prior to hybridization to
improve tissue penetration.
Probes will be prepared by various enzymatic procedures with a
reaction mixture that includes labelled nucleotide analogs or radioactive
nucleotides, or by direct synthesis as an oligonucleotide. Probes may carry
radioactive or fluorescent labels for direct detection or hapten labels for
detection by various indirect methods.
Once the sample has been prepared, it is incubated with the probe
at elevated temperature to allow the probe to hybridize to the sequence of
interest. Unhybridized probe is washed away and the remaining labelled probe is
detected.
Please contact us for any special needs in locating special gene
on chromosomes or in tissues by using ISH.
