Applications of Nanobodies in Biological Imaging

Background: Nanobodies (Nbs), derived from Camelidae heavy-chain antibodies, are single-domain fragments (15 kDa) with high antigen-binding specificity, enhanced tissue penetration, and low immunogenicity. These attributes address limitations of conventional antibodies, positioning Nbs as pivotal tools for targeted molecular imaging in diagnostics and therapeutics. Methods: Nbs are screened through phage/mRNA display or single B-cell sequencing, expressed in prokaryotic or yeast systems, and humanized via CDR grafting. Functional probes are engineered by conjugating Nbs with radionuclides (68Ga, 99mTc) or fluorophores (IRDye 800CW) for compatibility with PET, SPECT, NIRF, and ultrasound modalities. Results: Clinical trials validated Nb efficacy: 68Ga-HER2-Nb PET/CT achieved tumor-specific uptake in HER2+ cancers (NCT04467515), while 99mTc-PD-L1-Nb enabled quantitative SPECT-guided immunotherapy in NSCLC. NIRF-Nb conjugates (e.g., 11A4-800CW) enhanced intraoperative tumor delineation in murine models. Dual-targeted ultrasound microbubbles demonstrated multi-biomarker imaging via acoustic pressure modulation. Conclusion: Nbs advance biological imaging through superior resolution and rapid pharmacokinetics. Challenges persist in optimizing probe stability, minimizing immunogenicity, and scaling production. Future priorities include integrating multi-modal platforms, expanding applications to neurodegenerative disorders, and refining personalized diagnostic paradigms, underscoring their transformative potential in precision medicine.