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Against the backdrop of global pandemics caused by highly infectious diseases such as COVID-19 and Ebola virus disease (EVD), the development of innovative prevention and treatment strategies has become a strategic priority in the field of biomedicine. Neutralizing antibodies, as biological agents, are increasingly recognized for their potential in infectious disease control. Among these, nanobodies (Nbs) derived from camelid heavy-chain antibodies exhibit remarkable technical advantages due to their unique structural features. Compared to traditional neutralizing antibodies, nanobodies offer significant cost-effectiveness in production and enable versatile administration routes (e.g., subcutaneous injection, oral delivery, or aerosol inhalation), making them particularly suitable for respiratory infection control and resource-limited settings. Furthermore, engineered modification strategies\u2014including multivalent constructs, multi-epitope recognition designs, and fragment crystallizable (Fc) domain fusion\u2014effectively enhance their neutralizing activity and suppress viral immune escape mechanisms. Breakthroughs have been achieved in combating pathogens such as the Ebola virus and SARS-CoV-2, with mechanisms involving the blockade of virus\u2013host interactions, induction of viral particle disintegration, and enhancement of immune responses. This review comprehensively discusses the structural characteristics, high-throughput screening technologies, and engineering strategies of nanobodies, providing theoretical foundations for the development of novel antiviral therapeutics. These advances hold strategic significance for addressing emerging and re-emerging infectious diseases.<\/jats:p>","DOI":"10.3390\/microorganisms13061352","type":"journal-article","created":{"date-parts":[[2025,6,11]],"date-time":"2025-06-11T03:44:58Z","timestamp":1749613498000},"page":"1352","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Research Progress on the Application of Neutralizing Nanobodies in the Prevention and Treatment of Viral Infections"],"prefix":"10.3390","volume":"13","author":[{"given":"Qingling","family":"Duan","sequence":"first","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"},{"name":"Joint Programme of Nanchang University and Queen Mary University of London, Jiangxi Medicine School, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tong","family":"Ai","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yingying","family":"Ma","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ruoyu","family":"Li","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hanlin","family":"Jin","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xingyi","family":"Chen","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-6148-2843","authenticated-orcid":false,"given":"Rui","family":"Zhang","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kunlu","family":"Bao","sequence":"additional","affiliation":[{"name":"Department of Developmental BioEngineering, University of Twente, 7522 NB Enschede, The Netherlands"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qi","family":"Chen","sequence":"additional","affiliation":[{"name":"National Engineering Research Center for Bioengineering Drugs and the Technologies, Jiangxi Provincial Key Laboratory of Bioengineering Drugs, Institute of Translational Medicine, Jiangxi Medical College, Nanchang University, Nanchang 330031, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,6,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"495","DOI":"10.1038\/256495a0","article-title":"Continuous cultures of fused cells secreting antibody of predefined specificity","volume":"256","author":"Milstein","year":"1975","journal-title":"Nature"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Lu, R.-M., Hwang, Y.-C., Liu, I.-J., Lee, C.-C., Tsai, H.-Z., Li, H.-J., and Wu, H.-C. 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