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Mammalian cell display with automated oligo design and library assembly allows for rapid residue level conformational epitope mapping

Thalén, Niklas Berndt ; Karlander, Maximilian ; Lundqvist, Magnus ; Persson, Helena ; Hofström, Camilla ; Turunen, S. Pauliina ; Godzwon, Magdalena LU ; Volk, Anna Luisa ; Malm, Magdalena and Ohlin, Mats LU orcid , et al. (2024) In Communications Biology 7(1).
Abstract

Precise epitope determination of therapeutic antibodies is of great value as it allows for further comprehension of mechanism of action, therapeutic responsiveness prediction, avoidance of unwanted cross reactivity, and vaccine design. The golden standard for discontinuous epitope determination is the laborious X-ray crystallography method. Here, we present a combinatorial method for rapid mapping of discontinuous epitopes by mammalian antigen display, eliminating the need for protein expression and purification. The method is facilitated by automated workflows and tailored software for antigen analysis and oligonucleotide design. These oligos are used in automated mutagenesis to generate an antigen receptor library displayed on... (More)

Precise epitope determination of therapeutic antibodies is of great value as it allows for further comprehension of mechanism of action, therapeutic responsiveness prediction, avoidance of unwanted cross reactivity, and vaccine design. The golden standard for discontinuous epitope determination is the laborious X-ray crystallography method. Here, we present a combinatorial method for rapid mapping of discontinuous epitopes by mammalian antigen display, eliminating the need for protein expression and purification. The method is facilitated by automated workflows and tailored software for antigen analysis and oligonucleotide design. These oligos are used in automated mutagenesis to generate an antigen receptor library displayed on mammalian cells for direct binding analysis by flow cytometry. Through automated analysis of 33930 primers an optimized single condition cloning reaction was defined allowing for mutation of all surface-exposed residues of the receptor binding domain of SARS-CoV-2. All variants were functionally expressed, and two reference binders validated the method. Furthermore, epitopes of three novel therapeutic antibodies were successfully determined followed by evaluation of binding also towards SARS-CoV-2 Omicron BA.2. We find the method to be highly relevant for rapid construction of antigen libraries and determination of antibody epitopes, especially for the development of therapeutic interventions against novel pathogens.

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organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Communications Biology
volume
7
issue
1
article number
805
publisher
Nature Publishing Group
external identifiers
  • pmid:38961245
  • scopus:85197485847
ISSN
2399-3642
DOI
10.1038/s42003-024-06508-8
language
English
LU publication?
yes
id
16f2c08b-fa35-4ea9-ae0e-0088f705bdec
date added to LUP
2024-08-30 12:55:34
date last changed
2024-11-08 22:05:29
@article{16f2c08b-fa35-4ea9-ae0e-0088f705bdec,
  abstract     = {{<p>Precise epitope determination of therapeutic antibodies is of great value as it allows for further comprehension of mechanism of action, therapeutic responsiveness prediction, avoidance of unwanted cross reactivity, and vaccine design. The golden standard for discontinuous epitope determination is the laborious X-ray crystallography method. Here, we present a combinatorial method for rapid mapping of discontinuous epitopes by mammalian antigen display, eliminating the need for protein expression and purification. The method is facilitated by automated workflows and tailored software for antigen analysis and oligonucleotide design. These oligos are used in automated mutagenesis to generate an antigen receptor library displayed on mammalian cells for direct binding analysis by flow cytometry. Through automated analysis of 33930 primers an optimized single condition cloning reaction was defined allowing for mutation of all surface-exposed residues of the receptor binding domain of SARS-CoV-2. All variants were functionally expressed, and two reference binders validated the method. Furthermore, epitopes of three novel therapeutic antibodies were successfully determined followed by evaluation of binding also towards SARS-CoV-2 Omicron BA.2. We find the method to be highly relevant for rapid construction of antigen libraries and determination of antibody epitopes, especially for the development of therapeutic interventions against novel pathogens.</p>}},
  author       = {{Thalén, Niklas Berndt and Karlander, Maximilian and Lundqvist, Magnus and Persson, Helena and Hofström, Camilla and Turunen, S. Pauliina and Godzwon, Magdalena and Volk, Anna Luisa and Malm, Magdalena and Ohlin, Mats and Rockberg, Johan}},
  issn         = {{2399-3642}},
  language     = {{eng}},
  number       = {{1}},
  publisher    = {{Nature Publishing Group}},
  series       = {{Communications Biology}},
  title        = {{Mammalian cell display with automated oligo design and library assembly allows for rapid residue level conformational epitope mapping}},
  url          = {{http://dx.doi.org/10.1038/s42003-024-06508-8}},
  doi          = {{10.1038/s42003-024-06508-8}},
  volume       = {{7}},
  year         = {{2024}},
}