TY - JOUR
T1 - Single-domain near-infrared protein provides a scaffold for antigen-dependent fluorescent nanobodies
AU - Oliinyk, Olena S.
AU - Baloban, Mikhail
AU - Clark, Charles L.
AU - Carey, Erin
AU - Pletnev, Sergei
AU - Nimmerjahn, Axel
AU - Verkhusha, Vladislav V.
N1 - Funding Information:
We thank N. Peitsaro and N. Aarnio from the Flow Cytometry Core Facility and staff of the Neuronal Cell Culture Unit of the University of Helsinki for the technical assistance. This work was supported by grant nos. NS108034, NS112959, NS103522 and GM122567 from the US National Institutes of Health (NIH), grant no. 322226 from the Academy of Finland and the Magnus Ehrnrooth Foundation. S.P. was supported in part by the NIH Intramural Research Program for the Vaccine Research Center of the National Institute of Allergy and Infectious Diseases.
Publisher Copyright:
© 2022, The Author(s), under exclusive licence to Springer Nature America, Inc.
PY - 2022/6
Y1 - 2022/6
N2 - Small near-infrared (NIR) fluorescent proteins (FPs) are much needed as protein tags for imaging applications. We developed a 17 kDa NIR FP, called miRFP670nano3, which brightly fluoresces in mammalian cells and enables deep-brain imaging. By exploring miRFP670nano3 as an internal tag, we engineered 32 kDa NIR fluorescent nanobodies, termed NIR-Fbs, whose stability and fluorescence strongly depend on the presence of specific intracellular antigens. NIR-Fbs allowed background-free visualization of endogenous proteins, detection of viral antigens, labeling of cells expressing target molecules and identification of double-positive cell populations with bispecific NIR-Fbs against two antigens. Applying NIR-Fbs as destabilizing fusion partners, we developed molecular tools for directed degradation of targeted proteins, controllable protein expression and modulation of enzymatic activities. Altogether, NIR-Fbs enable the detection and manipulation of a variety of cellular processes based on the intracellular protein profile.
AB - Small near-infrared (NIR) fluorescent proteins (FPs) are much needed as protein tags for imaging applications. We developed a 17 kDa NIR FP, called miRFP670nano3, which brightly fluoresces in mammalian cells and enables deep-brain imaging. By exploring miRFP670nano3 as an internal tag, we engineered 32 kDa NIR fluorescent nanobodies, termed NIR-Fbs, whose stability and fluorescence strongly depend on the presence of specific intracellular antigens. NIR-Fbs allowed background-free visualization of endogenous proteins, detection of viral antigens, labeling of cells expressing target molecules and identification of double-positive cell populations with bispecific NIR-Fbs against two antigens. Applying NIR-Fbs as destabilizing fusion partners, we developed molecular tools for directed degradation of targeted proteins, controllable protein expression and modulation of enzymatic activities. Altogether, NIR-Fbs enable the detection and manipulation of a variety of cellular processes based on the intracellular protein profile.
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U2 - 10.1038/s41592-022-01467-6
DO - 10.1038/s41592-022-01467-6
M3 - Article
C2 - 35606446
AN - SCOPUS:85130437154
VL - 19
SP - 740
EP - 750
JO - Nature Methods
JF - Nature Methods
SN - 1548-7091
IS - 6
ER -