Epitope-specific T-cell Cloning
Epitope-specific T-cell Cloning - Related Videos
13:58
Peptide:MHC Tetramer-based Enrichment of Epitope-specific T cells
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Cited by 28 •
2012
This protocol describes the use of peptide:MHC tetramers and magnetic microbeads to isolate low frequency populations of epitope-specific T cells and analyze them by flow cytometry. This method enables the direct study of endogenous T cell populations of interest from in vivo experimental systems.
13:41
Use of Interferon-γ Enzyme-linked Immunospot Assay to Characterize Novel T-cell Epitopes of Human Papillomavirus
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Cited by 8 •
2012
Characterizing T-cell epitopes of pathogens that cause localized infections such as human papillomavirus is a challenge because of limited number of T cells in circulation. A method is described in which rare T cells were isolated and were characterized starting with a very small number of cells.
11:37
Zika Virus Specific Diagnostic Epitope Discovery
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Cited by 11 •
2017
In this protocol, we describe a technique to discover Zika virus specific diagnostic peptides using a high-density peptide microarray. This protocol can readly be adapted for other emerging infectious diseases.
09:55
Molecular Cloning
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2023
Molecular cloning is a set of methods, which are used to insert recombinant DNA into a vector - a carrier of DNA molecules that will replicate recombinant DNA fragments in host organisms. The DNA fragment, which may be a gene, can be isolated from a prokaryotic or eukaryotic specimen. Following isolation of the fragment of interest, or insert, both the vector and insert must be cut with restriction enzymes and purified. The purified pieces are joined together though a technique called...
05:21
Culturing and Harvesting of HSPC Clones: A Method to Obtain the Singly-Sorted HSPC Clones
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2023
This video describes the protocol for isolating HSPCs from the bone marrow and sorting them using a fluorescent activated cell sorting technique, which is a method to obtain singly-sorted HSPCs. We culture these sorted cells to obtain HSPC clones.
FAQs
The selected peptide or protein supplies antigenic specificity, while presentation by major histocompatibility complex molecules determines whether responding lymphocytes are stimulated. Consequently, the procedure enriches populations capable of recognizing the presented epitope in that presentation context rather than treating all T cells as equivalent. This principle matters when interpreting pathogen-specific recognition and other epitope-focused immune measurements.
Limiting dilution or another single-cell expansion method separates responding lymphocytes before expansion, allowing descendants of one starting cell to be analyzed as a clone. The resulting population can be tested for consistent recognition and functional behavior, rather than averaging responses from many unrelated cells. This distinction supports precise comparison of epitope-specific immune responses.
Individual clones allow researchers to connect defined antigen recognition with measurable immune functions, including cytokine production and cytotoxicity. Testing these properties separately can reveal whether recognition is accompanied by particular effector activities. Such functional profiles help characterize adaptive immune responses more precisely than recognition measurements alone, especially in studies of pathogen-specific immunity.
Cross-reactivity analysis examines whether a cloned population responds only to the selected epitope or also shows recognition of other tested antigenic targets. Because each clone can be evaluated separately, differences in recognition are not obscured by mixed lymphocyte populations. This information helps interpret the breadth and specificity of pathogen-related immune responses and complements epitope mapping.
A practical workflow begins by exposing lymphocytes to an antigenic peptide or protein presented by major histocompatibility complex molecules. Responding cells are then subjected to limiting dilution or another single-cell expansion approach, followed by growth of individual populations. Researchers can subsequently examine each clone for recognition, cytokine production, cytotoxicity, or cross-reactivity, linking isolation to measurable immune functions.
Researchers apply these clones when they need focused information about pathogen-specific or treatment-related T-cell responses. In infection studies, they can support epitope mapping and immune monitoring; in vaccine research, they help evaluate vaccine-induced responses. The same strategy can also examine therapeutic T-cell responses, providing a defined cellular system for comparing recognition and function.