Design and Construction of a Synthetic Nanobody Library: Testing Its Potential with a Single Selection Round Strategy

Area of Science:

• Biotechnology
• Immunology
• Molecular Biology

Background:

• Nanobodies (Nbs) are single-domain antibody fragments from Camelidae.
• Advantages include small size, high stability, and low production costs.
• Synthetic Nb libraries offer an alternative to animal immunization for generating antigen-specific Nbs.

Purpose of the Study:

• To design and construct a new synthetic nanobody library using phage display.
• To implement a structure-based approach with position-specific randomization of hypervariable loops.
• To evaluate the library's capability in selecting antigen-specific Nbs.

Main Methods:

• Construction of a synthetic nanobody library with 10^8 clonal diversity.
• Structure-based design with position-specific randomization of hypervariable loops.
• Phage display technology for library construction and Nb selection.

Main Results:

• The library exhibits high amino acid variability matching designed codon distributions.
• Successful selection of nanobodies specific for VEGF, TNF, and Andes virus GnGc complex.
• A single round of stringent biopanning yielded multiple antigen binders for each target.

Conclusions:

• The newly constructed synthetic nanobody library is a potent source for generating diverse antigen-specific Nbs.
• The library's design and selection strategy are effective for various applications.
• This approach offers a promising alternative for antibody discovery.