The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E
The commonly used antibodies 3D12 and 4D12 recognise the human leukocyte antigen E (HLA-E) protein. These antibodies bind distinct epitopes on HLA-E and differ in their ability to bind alleles of the major histocompatibility complex E (MHC-E) proteins of rhesus and cynomolgus macaques. We confirmed...
| Main Authors: | , , , , |
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| Format: | Journal article |
| Language: | English |
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Frontiers Media
2024
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| _version_ | 1826313243505721344 |
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| author | Brackenridge, S John, N Früh, K Borrow, P McMichael, A |
| author_facet | Brackenridge, S John, N Früh, K Borrow, P McMichael, A |
| author_sort | Brackenridge, S |
| collection | OXFORD |
| description | The commonly used antibodies 3D12 and 4D12 recognise the human leukocyte antigen E (HLA-E) protein. These antibodies bind distinct epitopes on HLA-E and differ in their ability to bind alleles of the major histocompatibility complex E (MHC-E) proteins of rhesus and cynomolgus macaques. We confirmed that neither antibody cross-reacts with classical HLA alleles, and used hybrids of different MHC-E alleles to map the regions that are critical for their binding. 3D12 recognises a region on the alpha 3 domain, with its specificity for HLA-E resulting from the amino acids present at three key positions (219, 223 and 224) that are unique to HLA-E, while 4D12 binds to the start of the alpha 2 domain, adjacent to the C terminus of the presented peptide. 3D12 staining is increased by incubation of cells at 27°C, and by addition of the canonical signal sequence peptide presented by HLA-E peptide (VL9, VMAPRTLVL). This suggests that 3D12 may bind peptide-free forms of HLA-E, which would be expected to accumulate at the cell surface when cells are incubated at lower temperatures, as well as HLA-E with peptide. Therefore, additional studies are required to determine exactly what forms of HLA-E can be recognised by 3D12. In contrast, while staining with 4D12 was also increased when cells were incubated at 27°C, it was decreased when the VL9 peptide was added. We conclude that 4D12 preferentially binds to peptidefree HLA-E, and, although not suitable for measuring the total cell surface levels of MHC-E, may putatively identify peptide-receptive forms. |
| first_indexed | 2024-03-07T08:27:09Z |
| format | Journal article |
| id | oxford-uuid:17e67e12-dcd8-4750-937e-60fb94c2257c |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-09-25T04:10:00Z |
| publishDate | 2024 |
| publisher | Frontiers Media |
| record_format | dspace |
| spelling | oxford-uuid:17e67e12-dcd8-4750-937e-60fb94c2257c2024-06-18T10:49:23ZThe antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-EJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:17e67e12-dcd8-4750-937e-60fb94c2257cEnglishSymplectic ElementsFrontiers Media2024Brackenridge, SJohn, NFrüh, KBorrow, PMcMichael, AThe commonly used antibodies 3D12 and 4D12 recognise the human leukocyte antigen E (HLA-E) protein. These antibodies bind distinct epitopes on HLA-E and differ in their ability to bind alleles of the major histocompatibility complex E (MHC-E) proteins of rhesus and cynomolgus macaques. We confirmed that neither antibody cross-reacts with classical HLA alleles, and used hybrids of different MHC-E alleles to map the regions that are critical for their binding. 3D12 recognises a region on the alpha 3 domain, with its specificity for HLA-E resulting from the amino acids present at three key positions (219, 223 and 224) that are unique to HLA-E, while 4D12 binds to the start of the alpha 2 domain, adjacent to the C terminus of the presented peptide. 3D12 staining is increased by incubation of cells at 27°C, and by addition of the canonical signal sequence peptide presented by HLA-E peptide (VL9, VMAPRTLVL). This suggests that 3D12 may bind peptide-free forms of HLA-E, which would be expected to accumulate at the cell surface when cells are incubated at lower temperatures, as well as HLA-E with peptide. Therefore, additional studies are required to determine exactly what forms of HLA-E can be recognised by 3D12. In contrast, while staining with 4D12 was also increased when cells were incubated at 27°C, it was decreased when the VL9 peptide was added. We conclude that 4D12 preferentially binds to peptidefree HLA-E, and, although not suitable for measuring the total cell surface levels of MHC-E, may putatively identify peptide-receptive forms. |
| spellingShingle | Brackenridge, S John, N Früh, K Borrow, P McMichael, A The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E |
| title | The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E |
| title_full | The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E |
| title_fullStr | The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E |
| title_full_unstemmed | The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E |
| title_short | The antibodies 3D12 and 4D12 recognise distinct epitopes and conformations of HLA-E |
| title_sort | antibodies 3d12 and 4d12 recognise distinct epitopes and conformations of hla e |
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