Immunofluorescence Detection
Immunofluorescence Detection - Related Videos
08:07
Simple Detection of Primary Cilia by Immunofluorescence
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Cited by 5 •
2020
Primary cilia are extracellular structures associated with the centriole. Primary cilia detection by immunofluorescent staining is a relatively simple procedure that results in extremely high-quality images. In this protocol, fibroblasts expressing primary cilia were fixed, immunostained, and imaged in a fluorescent or confocal microscope.
02:53
Immunofluorescence Assay to Detect Hepatitis C Virus Replication Complexes in Infected Cells
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2026
Source: Ren, S., et al. A Protocol for Analyzing Hepatitis C Virus Replication. J. Vis. Exp. (2014).This video demonstrates an immunofluorescence-based method to detect Hepatitis C virus replication in infected hepatoma cells. The protocol involves cell fixation, antibody staining for viral protein and double-stranded RNA, and visualization using fluorophore-tagged secondary antibodies. Dual signal detection under a fluorescence microscope confirms active viral replication within the host cells.
04:09
An Immunofluorescence Assay to Detect Alpha-Synuclein Aggregates in a Skin Biopsy Section
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2025
This video demonstrates an immunofluorescence assay to detect neurodegenerative disease-specific alpha-synuclein aggregates on skin biopsy sections. Free-floating immunostaining is employed to fluorescently label the carboxyl-terminal hydrolase and the alpha-synuclein aggregates in the dermal neurons. Under a confocal microscope, cells with fluorescence from both antibodies indicate the presence of alpha-synuclein aggregates in the dermal neurons.
04:18
An Immunofluorescence Staining Technique to Detect Adult Hippocampal Neurogenesis
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2025
The video illustrates an immunofluorescence staining method used to examine neurogenesis in the adult hippocampus. Initially, a mouse brain section undergoes treatment with primary antibodies directed at marker proteins associated with neurogenesis in neuronal progenitor cells. Next, it is labeled with secondary antibodies tagged with fluorophores. The brain section is then observed under a confocal microscope to distinguish various subtypes of progenitor cells, based on the expression patterns...
09:56
Immunofluorescence Microscopy: Immunofluorescence Staining of Paraffin-Embedded Tissue Sections
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2023
Source: Thomas Chaffee1, Thomas S. Griffith2,3,4, and Kathryn L. Schwertfeger1,3,4 1 Department of Lab Medicine and Pathology, University of Minnesota, Minneapolis, MN 55455 2 Department of Urology, University of Minnesota, Minneapolis, MN 55455 3 Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455 4 Center for Immunology, University of Minnesota, Minneapolis, MN 55455 Pathologic analyses of tissue sections can be used to obtain a better understanding of normal tissue...
FAQs
Direct immunofluorescence uses a fluorescently labeled primary antibody to recognize the target. In indirect immunofluorescence, an unlabeled primary antibody binds the target first, followed by a fluorophore-conjugated secondary antibody that binds the primary antibody. The distinction determines which antibody carries the detectable fluorophore and provides two ways to visualize antigen distribution in cells or tissues.
A fluorophore produces detectable light at a characteristic wavelength when excited by a microscope. Matching the microscope’s excitation and detection settings to that fluorophore allows the labeled target to be visualized. When several fluorophores are used, their distinct light characteristics help researchers distinguish multiple proteins or structures within the same specimen.
Specificity comes from the primary antibody recognizing its corresponding antigen in the sample. The fluorescent label then marks the location of that antibody-target interaction, allowing researchers to examine where a protein or structure occurs within cells or tissue. Accurate localization therefore depends on the antibody identifying the intended target rather than unrelated sample components.
A basic workflow begins with a biological sample containing the cells, tissue, or structures of interest. Researchers apply a primary antibody against the selected target, then either detect its fluorescent label directly or add a fluorophore-conjugated secondary antibody. A microscope excites the fluorophore and records the resulting light to reveal target distribution.
Multicolor immunofluorescence detection is useful when researchers need to examine several targets in one specimen rather than analyze each target separately. Different fluorophores provide distinguishable signals, supporting comparisons among proteins, cells, or structures within the same tissue or cultured-cell preparation. This approach helps relate molecular localization to cellular organization and experimental changes.
The method supports investigations of protein localization and cellular organization in tissue or cultured cells. It can also contribute to studies of disease mechanisms, diagnostics, and responses to experimental treatments. By showing where selected targets occur, alone or in combination with other labeled targets, the technique connects molecular observations with broader biological changes.