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Related Concept Videos

In-situ Hybridization02:31

In-situ Hybridization

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In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled probe. The technique was first used in 1969 for the investigation of nucleic acids. It is currently an essential tool in scientific research and clinical settings, especially for diagnostic purposes.
Types of probes and labels
A probe is a complementary strand of DNA or RNA that binds to corresponding nucleotide sequences in a cell. Many...
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FISH - Fluorescent In-situ Hybridization02:07

FISH - Fluorescent In-situ Hybridization

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Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...
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Related Experiment Video

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Whole Mount RNA Fluorescent in situ Hybridization of Drosophila Embryos
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Simultaneous Visualization of RNA and Protein Expression in Tissue Using a Combined RNAscope™ In Situ Hybridization

Anushka Dikshit1, Hailing Zong1, Courtney Anderson1

  • 1Advanced Cell Diagnostics, A Bio-Techne Brand, Newark, CA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|May 13, 2020
PubMed
Summary

This study combines RNAscope in situ hybridization (ISH) with immunofluorescence (IF) to visualize both gene and protein expression simultaneously. This dual-labeling approach provides spatial context for cellular analysis in complex tissues like tumors.

Keywords:
Dual ISH-IHCHeterogeneous tissueIn situ hybridizationRNAscopeSpatial profiling

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Gene expression analysis is crucial for understanding tissue complexity and disease mechanisms.
  • Traditional methods like PCR and sequencing lack spatial information, limiting analysis of heterogeneous tissues.
  • In situ hybridization (ISH) technologies, such as RNAscope, offer single-cell resolution for gene expression visualization.

Purpose of the Study:

  • To develop a protocol for simultaneous visualization of gene and protein expression within intact tissue samples.
  • To enable the study of cellular crosstalk and spatial relationships in complex biological systems.
  • To combine the spatial gene expression analysis capabilities of RNAscope ISH with immunofluorescence (IF).

Main Methods:

  • Detailed protocol for combining RNAscope in situ hybridization (ISH) with immunofluorescence (IF).
  • Application of the dual-labeling protocol to human tumor tissue and mouse brain tissue.
  • Simultaneous detection and visualization of target RNA and protein expression at single-cell resolution.

Main Results:

  • Successful integration of RNAscope ISH and IF assays into a single, effective protocol.
  • Demonstrated ability to visualize both gene and protein expression concurrently in complex tissue architectures.
  • Enabled high-resolution spatial analysis of molecular markers within human and mouse tissues.

Conclusions:

  • The combined RNAscope ISH and IF protocol is a powerful tool for simultaneous gene and protein expression analysis.
  • This technique provides critical spatial context for understanding cellular interactions and tissue heterogeneity.
  • The method is applicable to diverse tissue types, including human tumors and mouse brain, advancing molecular pathology and neuroscience research.