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Related Concept Videos

Immunofluorescence Microscopy01:12

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A fluorescence microscope uses fluorescent chromophores called fluorochromes, which can absorb energy from a light source and then emit this energy as visible light. Fluorochromes include naturally fluorescent substances (such as chlorophylls) and fluorescent stains that are added to the specimen to create contrast. Dyes such as Texas red and FITC are examples of fluorochromes. Other examples include the nucleic acid dyes 4’,6’-diamidino-2-phenylindole (DAPI), and acridine orange.
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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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Immunocytochemistry (ICC) and immunohistochemistry (IHC) are techniques that use antibodies to check for specific proteins or antigens in a sample. The technique was first published by Albert Coons in 1941 to detect the presence of pneumococcal antigen in tissue sections from mice infected with Pneumococcus. Immunocytochemistry helps localization of proteins or antigens in individual cells like blood cells, stem cells, etc., while immunohistochemistry does the same for tissue samples.
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Updated: Oct 11, 2025

Indirect Immunofluorescence on Frozen Sections of Mouse Mammary Gland
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Fluorescent Immunohistochemistry.

Vanessa Moreno1, Elizabeth A Smith1, Sergio Piña-Oviedo2

  • 1Arkana Laboratories, Little Rock, AR, USA.

Methods in Molecular Biology (Clifton, N.J.)
|December 3, 2021
PubMed
Summary
This summary is machine-generated.

Immunofluorescence is a sensitive technique using fluorescent antibodies to detect antigens in research and clinics. This guide details direct and indirect immunofluorescence methods and tissue preparation for accurate imaging.

Keywords:
Direct immunofluorescenceFFPEFresh-frozenImmunofluorescenceImmunohistochemistryIndirect immunofluorescence

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Area of Science:

  • Immunochemistry
  • Cell Biology
  • Histology

Background:

  • Immunofluorescence is a key immunochemical technique for detecting specific antigens.
  • It offers superior sensitivity and signal amplification compared to immunohistochemistry.
  • Requires specialized fluorescence microscopy for sample analysis.

Purpose of the Study:

  • To explain the principles of direct and indirect immunofluorescence.
  • To provide guidance on preparing tissues for immunofluorescence labeling.
  • To cover both fresh-frozen and formalin-fixed, paraffin-embedded tissues.

Main Methods:

  • Detailed explanation of direct immunofluorescence principles.
  • Detailed explanation of indirect immunofluorescence principles.
  • Protocols for preparing fresh-frozen and FFPE tissues for immunofluorescence.

Main Results:

  • Established protocols for tissue preparation.
  • Demonstrated principles of direct and indirect immunofluorescence.
  • Highlighted the importance of proper sample preparation for accurate results.

Conclusions:

  • Immunofluorescence is a versatile and sensitive diagnostic and research tool.
  • Proper tissue preparation is crucial for successful immunofluorescence.
  • Understanding direct and indirect methods enhances experimental design.