Retroviral transduction of murine primary T lymphocytes Journal Article
| Authors: | Lee, J.; Sadelain, M.; Brentjens, R. |
| Article Title: | Retroviral transduction of murine primary T lymphocytes |
| Abstract: | In comparison to human T cells, efficient retroviral gene transfer and subsequent expansion of murine primary T cells is more difficult to achieve. Herein, we describe an optimized gene transfer protocol utilizing an ecotropic viral vector to transduce primary murine T cells activated with magnetic beads coated with agonistic anti-CD3 and CD28 antibodies. Activated T cells are subsequently centrifuged (spinoculated) on RetroNectin-coated tissue culture plates in the context of retroviral supernatant. Variables found to be critical to high gene transfer and subsequent efficient T cell expansion included CD3/CD28 magnetic bead to cell ratio, time from T cell activation to initial spinoculation, frequency of T cell spin-oculation, interleukin-2 concentration in the medium, and the initial purity of the T cell preparation. |
| Keywords: | genetics; flow cytometry; cd3 antigen; t lymphocyte; antigens, cd3; t-lymphocytes; mouse; animal; cytology; metabolism; animals; mice; genetic transduction; transduction, genetic; immunology; cd28 antigen; antigens, cd28; retrovirus; retroviridae |
| Journal Title: | Methods in Molecular Biology |
| Volume: | 506 |
| ISSN: | 1064-3745 |
| Publisher: | Humana Press Inc |
| Date Published: | 2009-01-01 |
| Start Page: | 83 |
| End Page: | 96 |
| Language: | English |
| PUBMED: | 19110621 |
| PROVIDER: | scopus |
| DOI: | 10.1007/978-1-59745-409-4_7 |
| PMCID: | PMC5003426 |
| DOI/URL: | |
| Notes: | Chapter 7 in "Genetic Modification of Hematopoietic Stem Cells: Methods and Protocols" (ISBN: 978-1-58829-980-2) --- Cited By (since 1996): 2 - Export Date: 30 November 2010 - Source: Scopus |
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