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Nanobodies as Versatile Tool for Multiscale Imaging Modalities.

ABSTRACT: Molecular imaging is constantly growing in different areas of preclinical biomedical research. Several imaging methods have been developed and are continuously updated for both in vivo and in vitro applications, in order to increase the information about the structure, localization and function of molecules involved in physiology and disease. Along with these progresses, there is a continuous need for improving labeling strategies. In the last decades, the single domain antigen-binding fragments nanobodies (Nbs) emerged as important molecular imaging probes. Indeed, their small size (~15 kDa), high stability, affinity and modularity represent desirable features for imaging applications, providing higher tissue penetration, rapid targeting, increased spatial resolution and fast clearance. Accordingly, several Nb-based probes have been generated and applied to a variety of imaging modalities, ranging from in vivo and in vitro preclinical imaging to super-resolution microscopy. In this review, we will provide an overview of the state-of-the-art regarding the use of Nbs in several imaging modalities, underlining their extreme versatility and their enormous potential in targeting molecules and cells of interest in both preclinical and clinical studies.

SUBMITTER: Erreni M

PROVIDER: S-EPMC7767244 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Nanobodies as Versatile Tool for Multiscale Imaging Modalities.

Erreni Marco M Schorn Tilo T D'Autilia Francesca F Doni Andrea A

Biomolecules 20201218 12

Molecular imaging is constantly growing in different areas of preclinical biomedical research. Several imaging methods have been developed and are continuously updated for both in vivo and in vitro applications, in order to increase the information about the structure, localization and function of molecules involved in physiology and disease. Along with these progresses, there is a continuous need for improving labeling strategies. In the last decades, the single domain antigen-binding fragments ...[more]

PMID: 33353213

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Project description:There is a growing need for a highly stable system to allow the production of biologics for diagnoses and therapeutic interventions on demand that could be used in extreme environments. Among the variety of biologics, nanobodies (Nbs) derived from single-chain variable antibody fragments from camelids have attracted great attention in recent years due to their small size and great stability with translational potentials in whole-body imaging and the development of new drugs. Intracellular expression using the bacterium Escherichia coli has been the predominant system to produce Nbs, and this requires lengthy steps for releasing intracellular proteins for purification as well as removal of endotoxins. Lyophilized, translationally competent cell extracts have also been explored as offering portability and long shelf life, but such extracts may be difficult to scale up and suffer from batch-to-batch variability. To address these problems, we present a new system to do the following: (i) engineer the spore-forming bacterium Bacillus subtilis to secrete Nbs that can target small molecules or protein antigens on mammalian cells, (ii) immobilize Nbs containing a cellulose-binding domain on a cellulose matrix for long-term storage and small-molecule capturing, (iii) directly use Nb-containing bacterial supernatant fluid to perform protein detection on cell surfaces, and (iv) convert engineered B. subtilis to spores that are resistant to most environmental extremes. In summary, our work may open a new paradigm for using B. subtilis as an extremely stable microbial factory to produce Nbs with applications in extreme environments on demand.IMPORTANCE It is highly desirable to produce biologics for diagnoses and therapeutic interventions on demand that could be used in a variety of settings. Among the many biologics, Nbs have attracted attention due to their small size, thermal stability, and broad utility in diagnoses, therapies, and fundamental research. Nbs originate from antibodies found in camelids, and >10 companies have invested in Nbs as potential drugs. Here, we present a system using cells of the bacterium Bacillus subtilis as a versatile platform for production of Nbs and then antigen detection via customized affinity columns. Importantly, B. subtilis carrying engineered genes for Nbs can form spores, which survive for years in a desiccated state. However, upon rehydration and exposure to nutrients, spores rapidly transition to growing cells which secrete encoded Nbs, thus allowing their manufacture and purification.

Dataset Information

Nanobodies as Versatile Tool for Multiscale Imaging Modalities.

Publications

Nanobodies as Versatile Tool for Multiscale Imaging Modalities.

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