Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries.
The use of Xenopus laevis as a model for vertebrate developmental biology is limited by a lack of antibodies specific for embryonic antigens. This study evaluated the use of immune and non-immune phage display libraries for the isolation of single domain antibodies, or nanobodies, with specificities...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2019-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0216083 |
| _version_ | 1819291915686772736 |
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| author | Keiji Itoh Alice H Reis Andrew Hayhurst Sergei Y Sokol |
| author_facet | Keiji Itoh Alice H Reis Andrew Hayhurst Sergei Y Sokol |
| author_sort | Keiji Itoh |
| collection | DOAJ |
| description | The use of Xenopus laevis as a model for vertebrate developmental biology is limited by a lack of antibodies specific for embryonic antigens. This study evaluated the use of immune and non-immune phage display libraries for the isolation of single domain antibodies, or nanobodies, with specificities for Xenopus embryonic antigens. The immune nanobody library was derived from peripheral blood lymphocyte RNA obtained from a llama immunized with Xenopus gastrula homogenates. Screening this library by immunostaining of embryonic tissues with pooled periplasmic material and sib-selection led to the isolation of several monoclonal phages reactive with the cytoplasm and nuclei of gastrula cells. One antigen recognized by a group of nanobodies was identified using a reverse proteomics approach as nucleoplasmin, an abundant histone chaperone. As an alternative strategy, a semi-synthetic non-immune llama nanobody phage display library was panned on highly purified Xenopus proteins. This proof-of-principle approach isolated monoclonal nanobodies that specifically bind Nuclear distribution element-like 1 (Ndel1) in multiple immunoassays. Our results suggest that immune and non-immune phage display screens on crude and purified embryonic antigens can efficiently identify nanobodies useful to the Xenopus developmental biology community. |
| first_indexed | 2024-12-24T03:46:14Z |
| format | Article |
| id | doaj.art-07473f47543c4fd19b2c99767c96ade1 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-24T03:46:14Z |
| publishDate | 2019-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-07473f47543c4fd19b2c99767c96ade12022-12-21T17:16:42ZengPublic Library of Science (PLoS)PLoS ONE1932-62032019-01-01145e021608310.1371/journal.pone.0216083Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries.Keiji ItohAlice H ReisAndrew HayhurstSergei Y SokolThe use of Xenopus laevis as a model for vertebrate developmental biology is limited by a lack of antibodies specific for embryonic antigens. This study evaluated the use of immune and non-immune phage display libraries for the isolation of single domain antibodies, or nanobodies, with specificities for Xenopus embryonic antigens. The immune nanobody library was derived from peripheral blood lymphocyte RNA obtained from a llama immunized with Xenopus gastrula homogenates. Screening this library by immunostaining of embryonic tissues with pooled periplasmic material and sib-selection led to the isolation of several monoclonal phages reactive with the cytoplasm and nuclei of gastrula cells. One antigen recognized by a group of nanobodies was identified using a reverse proteomics approach as nucleoplasmin, an abundant histone chaperone. As an alternative strategy, a semi-synthetic non-immune llama nanobody phage display library was panned on highly purified Xenopus proteins. This proof-of-principle approach isolated monoclonal nanobodies that specifically bind Nuclear distribution element-like 1 (Ndel1) in multiple immunoassays. Our results suggest that immune and non-immune phage display screens on crude and purified embryonic antigens can efficiently identify nanobodies useful to the Xenopus developmental biology community.https://doi.org/10.1371/journal.pone.0216083 |
| spellingShingle | Keiji Itoh Alice H Reis Andrew Hayhurst Sergei Y Sokol Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries. PLoS ONE |
| title | Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries. |
| title_full | Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries. |
| title_fullStr | Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries. |
| title_full_unstemmed | Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries. |
| title_short | Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries. |
| title_sort | isolation of nanobodies against xenopus embryonic antigens using immune and non immune phage display libraries |
| url | https://doi.org/10.1371/journal.pone.0216083 |
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