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<p>Understanding the mechanistic interplay between antibodies and invading pathogens is essential for vaccine development. Current methods are labour and time intensive and limited by sample preparation bottlenecks. Here we present microfluidic electron microscopy-based polyclonal epitope mapping (mEM), which combines microfluidics with single-particle electron microscopy for the structural characterization of immune complexes using small volumes of sera (&lt;4 µl). First, we used mEM to map polyclonal antibodies present in sera from infected and vaccinated individuals against five viral glycoproteins using negative-stain electron microscopy. The mEM detected a greater number of epitopes compared with conventional polyclonal epitope structural mapping methods. Second, we used mEM and cryo-electron microscopy to characterize two coronavirus spikes and one HA glycoprotein with and without polyclonal antibodies. Finally, we mapped individual antibody responses over time in mice vaccinated with human immunodeficiency virus envelope N332-GT5. mEM enables the rapid, high-throughput mapping of antibodies targeting a broad range of glycoproteins, facilitating a better understanding of infection and guiding structure-based vaccine design.</p>

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Microfluidics combined with electron microscopy for rapid and high-throughput mapping of antibody–viral glycoprotein complexes

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年6月3日

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  • Leigh M. Sewall,
  • Rebeca de Paiva Froes Rocha,
  • Grace Gibson,
  • Michelle Louie,
  • Zhenfei Xie,
  • Sandhya Bangaru,
  • Andy S. Tran,
  • Gabriel Ozorowski,
  • Subhasis Mohanty,
  • Nathan Beutler,
  • Thomas F. Rogers,
  • Dennis R. Burton,
  • Albert C. Shaw,
  • Facundo D. Batista,
  • Blanca Chocarro Ruiz,
  • Alba Torrents de la Peña,
  • Andrew B. Ward

摘要

Understanding the mechanistic interplay between antibodies and invading pathogens is essential for vaccine development. Current methods are labour and time intensive and limited by sample preparation bottlenecks. Here we present microfluidic electron microscopy-based polyclonal epitope mapping (mEM), which combines microfluidics with single-particle electron microscopy for the structural characterization of immune complexes using small volumes of sera (<4 µl). First, we used mEM to map polyclonal antibodies present in sera from infected and vaccinated individuals against five viral glycoproteins using negative-stain electron microscopy. The mEM detected a greater number of epitopes compared with conventional polyclonal epitope structural mapping methods. Second, we used mEM and cryo-electron microscopy to characterize two coronavirus spikes and one HA glycoprotein with and without polyclonal antibodies. Finally, we mapped individual antibody responses over time in mice vaccinated with human immunodeficiency virus envelope N332-GT5. mEM enables the rapid, high-throughput mapping of antibodies targeting a broad range of glycoproteins, facilitating a better understanding of infection and guiding structure-based vaccine design.