Nanobody preparation and functional studies for broad⁃spectrum neutralization of SARS⁃ CoV⁃2 and its variants
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1Medical Research Center,Women’s Hospital of Nanjing Medical University(Nanjing Women and Children’sHealthcare Hospital),2Nanjing Medical Key Laboratory of Female Fertility Preservation and Restoration,Nanjing 210004 ,China

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R392

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    Abstract:

    Objective:To produce broad - spectrum nanobodies(Nbs)targeting severe acute respiratory syndrome coronavirus 2 (SARS - CoV - 2)and its variants,and establish a strategy for producing Nbs broad - spectrum neutralizing the virus and its various variants. Methods:Alpacas were immunized with the receptor-binding domain(RBD)of the SARS-CoV-2 wild-type strain(WT-RBD) to construct a nanobody phage library,and the library was screened by biopanning and ELISA with the RBD of the Delta variant(Delta- RBD)to select Nbs. The selected Nbs were expressed and purified using the Pichia pastoris system,and their properties and functions characterized by ELISA,biolayer interferometry,pseudovirus neutralization experiments,and molecular modeling and docking. Results:Four Nbs were obtained,all exibiting high affinity for WTRBD,while also effectively neutralizing DeltaRBD and the RBD of the Omicron variant(Omicron RBD),with Nb45 effectively neutralizing the pseudovirus of(Omicron -RBD). We also found that the mechanism of these Nbs for neutralizing SARSCoV-2 and its variants may be related to interfering with the conserved sites on these RBD. Conclusion:The four Nbs obtained have great potential for prevention and treatment of SARS-CoV-2 and its variants,and the strategy established in this study can also be widely used for the preparation of Nbs broad - spectrum neutralizing viruses and their various variants.

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WANG Baicheng, NI Jingyi, LIU Xinyue, GONG Xue, JI Chenbo. Nanobody preparation and functional studies for broad⁃spectrum neutralization of SARS⁃ CoV⁃2 and its variants[J].,2025,45(11):1598-1607.

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History
  • Received:January 21,2025
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  • Online: November 12,2025
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