Tcr Cloning
Tcr Cloning - Related Videos
19:05
Measuring TCR-pMHC Binding In Situ using a FRET-based Microscopy Assay
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Cited by 9 •
2015
This manuscript describes how to conduct (single molecule) Förster Resonance Energy Transfer (FRET)- based assays to measure the binding dynamics between T-cell antigen receptor (TCR) and antigenic peptide-loaded MHC molecules as they occur within the immunological synapse of a T-cell in contact with a functionalized planar supported lipid bilayer.
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...
11:21
Streamlined Single Cell TCR Isolation and Generation of Retroviral Vectors for In Vitro and In Vivo Expression of Human TCRs
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Cited by 3 •
2017
The current protocol combines single cell paired human TCR alpha and beta chain sequencing with streamlined generation of retroviral vectors compatible with in vitro and in vivo TCR expression.
01:27
Reproductive Cloning
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2019
Reproductive cloning is the process of producing a genetically identical copy—a clone—of an entire organism. While clones can be produced by splitting an early embryo—similar to what happens naturally with identical twins—cloning of adult animals is usually done by a process called somatic cell nuclear transfer (SCNT). Somatic Cell Nuclear Transfer In SCNT, an egg cell is taken from an animal and its nucleus is removed, creating an enucleated egg. Then a somatic cell—any cell that is not a sex...
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
Correct pairing preserves the receptor configuration associated with the original antigen-responsive T cell. After the recovered alpha and beta sequences are expressed together, the chains assemble at the recipient cell surface, allowing investigators to examine a defined antigen-recognition property rather than a mixed receptor population. This makes receptor-specific comparisons more interpretable.
These cells provide receptor sequences linked to a demonstrated response to an antigen. Recovering alpha and beta genes from that population connects the molecular sequence to antigen recognition observed in the source cells. The resulting clones can therefore support characterization of immune responses and comparison of receptor function in a more defined experimental system.
Introducing different cloned alpha and beta chain sequences into recipient cells creates cells with defined receptor specificities. Investigators can then compare the antigen-recognition properties associated with those receptors, rather than evaluating an unfractionated collection of T cells. In infection studies, this strategy helps distinguish receptor-level differences among responses directed toward pathogen-related antigens.
Researchers begin with antigen-responsive T cells, recover the genes encoding the TCR alpha and beta chains, and amplify those sequences. They then insert the amplified genes into an expression vector and introduce the construct into recipient cells. Successful chain assembly at the cell surface provides the basis for examining the receptor's defined antigen-recognition properties.
It can identify receptors associated with responses to pathogens and provide a way to characterize those immune responses at the receptor level. By expressing selected alpha and beta chains in recipient cells, researchers can examine defined antigen-recognition properties and compare receptors linked to different infection-related responses. This helps connect T-cell receptor sequence information with immunological function.
Applications include identifying pathogen-specific receptors, comparing receptor function, and characterizing immune responses. The technique also supports development of engineered T cells for studying or targeting infected cells. Because the expressed chains confer defined antigen-recognition properties, investigators can evaluate selected receptors in a cellular context and relate receptor choice to the intended infection-focused research question.