Detection and manipulation of live antigen-expressing cells using conditionally stable nanobodies
Tang, Jonathan CY
Ellis, Emily G
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CitationTang, Jonathan CY, Eugene Drokhlyansky, Behzad Etemad, Stephanie Rudolph, Binggege Guo, Sui Wang, Emily G Ellis, Jonathan Z Li, and Constance L Cepko. 2016. “Detection and manipulation of live antigen-expressing cells using conditionally stable nanobodies.” eLife 5 (1): e15312. doi:10.7554/eLife.15312. http://dx.doi.org/10.7554/eLife.15312.
AbstractThe ability to detect and/or manipulate specific cell populations based upon the presence of intracellular protein epitopes would enable many types of studies and applications. Protein binders such as nanobodies (Nbs) can target untagged proteins (antigens) in the intracellular environment. However, genetically expressed protein binders are stable regardless of antigen expression, complicating their use for applications that require cell-specificity. Here, we created a conditional system in which the stability of an Nb depends upon an antigen of interest. We identified Nb framework mutations that can be used to rapidly create destabilized Nbs. Fusion of destabilized Nbs to various proteins enabled applications in living cells, such as optogenetic control of neural activity in specific cell types in the mouse brain, and detection of HIV-infected human cells by flow cytometry. These approaches are generalizable to other protein binders, and enable the rapid generation of single-polypeptide sensors and effectors active in cells expressing specific intracellular epitopes. DOI: http://dx.doi.org/10.7554/eLife.15312.001
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