what are the unique features of nanobody and the applications
What are the unique features of nanobodies
Nanobody show some striking advantages versus conventional Abs that are noteworthy since they combine desirable features of mAbs with some of the beneficial properties of small molecule drugs.
- They can bind a broad range of epitopes showing affinities in the nanomolar or even picomolar range. This implies that the affinities of Nbs are comparable or superior to those of conventional mAbs, reaching picomolar affinity constant range.
- Possibility of formatting or multimerization. Given the small size and their monomeric nature of Nbs, they can be genetically engineered to obtain modular blocks that can be fused to form a new construct which will enable multispecificity and versatility.
- They are highly stable and soluble. Nbs present optimal biophysical and biochemical properties including solubility, thermal tolerance, and proteolytic resistance. The folding of CDR3 loop and the hydrophilic content of the framework-2 region gives them high solubility in aqueous solutions and lack of aggregation.
- Deep and fast tissue penetration and rapid blood clearance. Since passive intercellular diffusion rate within a tissue depends on the molecular size and is approximately inversely proportional to it, a 15-kDa monovalent Nb should improve the penetrability when compared with the performance of conventional mAbs (150 kDa).
- Recognition of hidden epitopes. Crystallographic studies of conventional Abs have revealed that in most cases the Ag-binding surface is flat or concave.
- Immunogenicity. Camelid VHH domains present high degree of homology with human VH domains, which means that to date they have not shown any unexpected immunogenicity reactions, which can be a potential concern for clinical applications
- Production costs. mAbs are large multimeric proteins that usually undergo post-translational modifications. Hence, their production requires sophisticated machinery only found in eukaryotic systems.
Applications of nanobodies as diagnostic and therapeutic tools
- Detection of proteins and microorganisms
- Detection of small molecules
- Imaging applications
- Therapeutic tools
Nanobody production scheme using a phage display library. The genetic information can be obtained by active immunization using an
immunogen or using non-immunized animals by collection of blood which contains the lymphocytes. Increasing the diversity using semisynthetic libraries and randomly introducing amino acids in the CDR region is possible in order to improve the binding capabilities.
After the insertion of the plasmid in a bacterophage, the phagemid library is ready for this selection. Phage ELISA is the usual way to perform the biopanning. After the best candidate is selected, the production of the nanobody can be expressed in bacteria, yeast, or mammalian cells through its corresponding expression vector
Ref: Nanobody: outstanding features for diagnostic and therapeutic application. Anal Bioanal Chem. 2019 Mar;411(9):1703-1713. doi: 10.1007 s00216-019-01633-4. Epub 2019 Feb 8.