Journal
PLOS ONE
Volume 8, Issue 3, Pages -Publisher
PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0060298
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Categories
Funding
- CNPq
- Instituto do MIlenio de Terapia Genica
- CAPES
- Fundacao do Cancer
- INCa
- Fundacao de Amparo a Pesquisa do Rio de Janeiro Carlos Chagas Filho
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Gene transfer to T lymphocytes has historically relied on retro and lentivirus, but recently transposon-based gene transfer is rising as a simpler and straight forward approach to achieve stable transgene expression. Transfer of expression cassettes to T lymphocytes remains challenging, being based mainly on commercial kits. Aims: We herein report a convenient and affordable method based on in house made buffers, generic cuvettes and utilization of the widely available Lonza nucleofector II device to promote efficient gene transfer to T lymphocytes. Results: This approach renders high transgene expression levels in primary human T lymphocytes (mean 45%, 41-59%), the hard to transfect murine T cells (mean 38%, 36-42% for C57/BL6 strain) and human Jurkat T cell line. Cell viability levels after electroporation allowed further manipulations such as in vitro expansion and Chimeric Antigen Receptor (CAR) mediated gain of function for target cell lysis. Conclusions: We describe here an efficient general protocol for electroporation based modification of T lymphocytes. By opening access to this protocol, we expect that efficient gene transfer to T lymphocytes, for transient or stable expression, may be achieved by an increased number of laboratories at lower and affordable costs.
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