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PLoS One. 2013;8(3):e60298. doi: 10.1371/journal.pone.0060298. Epub 2013 Mar 26.

An efficient low cost method for gene transfer to T lymphocytes.

Author information

1
Programa de Carcinogênese Molecular, Coordenação de Pesquisa (CPQ), Instituto Nacional de Câncer (INCA), Rio de Janeiro, Brazil.

Abstract

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.

PMID:
23555950
PMCID:
PMC3608570
DOI:
10.1371/journal.pone.0060298
[Indexed for MEDLINE]
Free PMC Article

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