Article ; Online: A Transgenic Heavy Chain IgG Mouse Platform as a Source of High Affinity Fully Human Single-Domain Antibodies for Therapeutic Applications.
Methods in molecular biology (Clifton, N.J.)
2022 Volume 2446, Page(s) 121–141
Abstract: The antibody repertoires of transgenic mice expressing human heavy chain only antibodies (HCAbs) can be retrieved from immune cells after antigen challenge. Compared with genetically modified rodents expressing conventional human antibodies (tetramers ... ...
| Abstract | The antibody repertoires of transgenic mice expressing human heavy chain only antibodies (HCAbs) can be retrieved from immune cells after antigen challenge. Compared with genetically modified rodents expressing conventional human antibodies (tetramers consisting of two heavy chains paired with two light chains), there is no chain pairing problem, since each antibody consists of a heavy chain dimer which is solely responsible for antigen binding. HCAbs can be obtained by classical hybridoma fusion, or the generation of phage libraries or eukaryotic cell libraries displaying or secreting HCAbs. Combined transcriptomic/serum proteomic approaches can also be used to determine the repertoire of antibodies, as well as single cell technologies such as the Beacon system that enable capture of immune cells of interest, analysis, and sequencing of antibodies in a short period of time. Here, we describe a protocol for obtaining monoclonal HCAbs from immunized Harbour transgenic mice through the generation and screening of HEK cell libraries of secreted antibodies. The method can be used routinely and is fast and affordable for everyone. Selected VH regions (single domains) are sequenced and individual HCAbs can be produced and purified from the same expression vector that is used for library generation (hIgG1 Fc). They can also be cloned into other expression plasmids and reformatted to equip them with a particular effector function, modify lifespan in serum, or optimize valency and avidity depending on the specific aim. |
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| MeSH term(s) | Animals ; Humans ; Immunoglobulin G ; Immunoglobulin Heavy Chains/genetics ; Mice ; Mice, Transgenic ; Proteomics ; Single-Domain Antibodies |
| Chemical Substances | Immunoglobulin G ; Immunoglobulin Heavy Chains ; Single-Domain Antibodies |
| Language | English |
| Publishing date | 2022-02-14 |
| Publishing country | United States |
| Document type | Journal Article |
| ISSN | 1940-6029 |
| ISSN (online) | 1940-6029 |
| DOI | 10.1007/978-1-0716-2075-5_6 |
| Database | MEDical Literature Analysis and Retrieval System OnLINE |
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