Isolation of nanobodies against Xenopus embryonic antigens using immune and non-immune phage display libraries.
Amino Acid Sequence
Animals
Antibodies
/ isolation & purification
Antigens
/ immunology
Cell Surface Display Techniques
/ methods
Cytoskeletal Proteins
/ immunology
Embryonic Development
/ immunology
Gastrula
Peptide Library
Single-Domain Antibodies
/ immunology
Stage-Specific Embryonic Antigens
/ metabolism
Xenopus Proteins
/ genetics
Xenopus laevis
/ embryology
Journal
PloS one
ISSN: 1932-6203
Titre abrégé: PLoS One
Pays: United States
ID NLM: 101285081
Informations de publication
Date de publication:
2019
2019
Historique:
received:
02
01
2019
accepted:
12
04
2019
entrez:
4
5
2019
pubmed:
3
5
2019
medline:
23
2
2020
Statut:
epublish
Résumé
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.
Identifiants
pubmed: 31048885
doi: 10.1371/journal.pone.0216083
pii: PONE-D-19-00108
pmc: PMC6497274
doi:
Substances chimiques
Antibodies
0
Antigens
0
Cytoskeletal Proteins
0
Ndel1 protein, Xenopus
0
Peptide Library
0
Single-Domain Antibodies
0
Stage-Specific Embryonic Antigens
0
Xenopus Proteins
0
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
e0216083Subventions
Organisme : NIGMS NIH HHS
ID : R01 GM116657
Pays : United States
Déclaration de conflit d'intérêts
The authors have declared that no competing interests exist.
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