Detection and manipulation of live antigen-expressing cells using conditionally stable nanobodies
- Howard Hughes Medical Institute, Harvard Medical School, United States
- Harvard Medical School, United States
Abstract
The 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 proteins.
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Ethics
Animal experimentation: The Institutional Animal Care and Use Committee at Harvard Medical School approved all animal experiments conducted under protocols 428-R98, 04537 and 1493.
Copyright
© 2016, Tang et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Detection and manipulation of live antigen-expressing cells using conditionally stable nanobodies
eLife 5:e15312.
https://doi.org/10.7554/eLife.15312
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