Genetic in vivo engineering of human T lymphocytes in mouse models.
Journal
Nature protocols
ISSN: 1750-2799
Titre abrégé: Nat Protoc
Pays: England
ID NLM: 101284307
Informations de publication
Date de publication:
07 2021
07 2021
Historique:
received:
23
09
2020
accepted:
21
01
2021
pubmed:
14
4
2021
medline:
21
7
2021
entrez:
13
4
2021
Statut:
ppublish
Résumé
Receptor targeting of vector particles is a key technology to enable cell type-specific in vivo gene delivery. For example, T cells in humanized mouse models can be modified by lentiviral vectors (LVs) targeted to human T-cell markers to enable them to express chimeric antigen receptors (CARs). Here, we provide detailed protocols for the generation of CD4- and CD8-targeted LVs (which takes ~9 d in total). We also describe how to humanize immunodeficient mice with hematopoietic stem cells (which takes 12-16 weeks) and precondition (over 5 d) and administer the vector stocks. Conversion of the targeted cell type is monitored by PCR and flow cytometry of blood samples. A few weeks after administration, ~10% of the targeted T-cell subtype can be expected to have converted to CAR T cells. By closely following the protocol, sufficient vector stock for the genetic manipulation of 10-15 humanized mice is obtained. We also discuss how the protocol can be easily adapted to use LVs targeted to other types of receptors and/or for delivery of other genes of interest.
Identifiants
pubmed: 33846629
doi: 10.1038/s41596-021-00510-8
pii: 10.1038/s41596-021-00510-8
doi:
Substances chimiques
Antigens, CD
0
Receptors, Chimeric Antigen
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
3210-3240Subventions
Organisme : NIAID NIH HHS
ID : R01 AI145045
Pays : United States
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