Elispot Method
Elispot Method - Related Videos
11:53
ELISPOT Assay: Detection of IFN-γ Secreting Splenocytes
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2023
Source: Tonya J. Webb1 1 Department of Microbiology and Immunology, University of Maryland School of Medicine and the Marlene and Stewart Greenebaum Comprehensive Cancer Center, Baltimore, Maryland 21201 ELISPOT is a standardized, reproducible assay used to detect cellular immune responses. The assay utilizes an enzyme-linked immunosorbent assay (ELISA)- based method to detect single-cell immune responses which can be visualized by spots, hence the name ELISPOT. ELISPOT was first described in...
04:20
Functional Assessment of Antibody-Secreting B Cells Using ELISpot Assay
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2025
This video demonstrates the use of the ELISpot assay to assess the function of antibody-secreting B cells, ASCs. The antibodies secreted by the ASCs interact with the membrane-bound polyclonal antibody fragments, which are detected by the enzyme-conjugated IgG or IgM antibodies. The colored spot produced in each well indicates the presence of the specific antibody's isotype, which correlates with the B cells' secretory activity.
08:26
The Isolation, Differentiation, and Quantification of Human Antibody-secreting B Cells from Blood: ELISpot as a Functional Readout of Humoral Immunity
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Cited by 10 •
2016
Human peripheral blood is commonly used for the assessment of the humoral immune response. Here, the methods for isolating human B cells from peripheral blood, differentiating human B cells into antibody (Ab)-secreting B cells (ASCs) in culture, and enumerating the total IgM- and IgG-ASCs via an ELISpot assay are described.
06:13
Enzyme-linked Immunospot Assay (ELISPOT): Quantification of Th-1 Cellular Immune Responses Against Microbial Antigens
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Cited by 3 •
2010
Identification of microbial targets of adaptive immunity in idiopathic diseases can be accomplished by the use of the enzyme-linked immunospot assay.
08:13
Optimized Interferon-gamma ELISpot Assay to Measure T Cell Responses in the Guinea Pig Model after Vaccination
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Cited by 3 •
2019
The development of the interferon-gamma ELISpot assay for guinea pig PBMCs allows the characterization of T-cell responses in this highly relevant model for studying infectious diseases. We have applied the assay to measure T cell responses associated with intradermal delivery of DNA vaccines.
FAQs
The membrane-bound capture antibody retains the secreted cytokine, antibody, or other target biomolecule close to the cell that released it. A labeled detection antibody then binds the immobilized material, producing a visible spot. This paired-antibody arrangement connects each signal to a producing cell and supports measurement of target-specific secretion.
Spot counts primarily indicate the frequency of cells responding by releasing the measured biomolecule. Spot size or intensity can add information about the amount or degree of secretion associated with those responses. Considering these features together helps distinguish how many cells responded from how strongly they appeared to secrete the target.
Because secreted molecules remain localized near the individual cells that produce them, the assay can reveal responses from cells that represent only a small fraction of the cultured population. This single-cell resolution provides information that would be obscured if all secreted material were considered only as a combined culture-level signal.
Cells are placed in culture on a membrane coated with an antibody that captures the selected secreted molecule. During culture, released material becomes immobilized near its source cell. A labeled detection antibody is then applied to identify the captured molecules, and the resulting visible spots are counted and evaluated.
Researchers apply it to measure antigen-specific T-cell and B-cell responses, evaluate vaccine immunogenicity, investigate pathogen exposure, and assess immune-cell function. These uses make the assay relevant when the central question concerns whether particular immune cells respond to an antigen or pathogen-associated stimulus through secretion.
In infection research, spot patterns can indicate the frequency of immune cells responding to a pathogen-related antigen and can provide additional secretion information through spot size or intensity. The same measurements support comparisons of immune-cell function, pathogen exposure, and vaccine-induced responses when the assay targets the relevant secreted biomolecule.