David B. Lewis

Publication Details

  • Consistent transient transfection of DNA into non-transformed human and murine T-lymphocytes JOURNAL OF IMMUNOLOGICAL METHODS Cron, R. Q., Schubert, L. A., Lewis, D. B., HUGHES, C. C. 1997; 205 (2): 145-150

    Abstract:

    The ability to analyze transcriptional regulation in non-transformed T-cells has been hampered by the inability to reproducibly transiently transfect these cells with DNA constructs. We have previously demonstrated that normal human whole mononuclear and CD4 T-cells can be consistently transiently transfected with plasmid DNA. Human cells were most receptive to plasmid DNA uptake between 19.5 and 20 h after prestimulation with a submitogenic dose of the polyclonal T-cell activator, PHA. Here we report an alteration and optimization of this protocol for non-transformed murine splenic T-cells, using concanavalin A instead of PHA as the preactivation stimulus. When coupled with the high sensitivity of luciferase reporter gene constructs, this protocol facilitates the analysis of a variety of T-cell-specific promoters in non-transformed T-cells. In addition, we directly demonstrate that murine T-cells are specifically transiently transfected among a population of whole mononuclear cells by using an expression vector for green fluorescent protein.

    View details for Web of Science ID A1997XR21700005

    View details for PubMedID 9294595

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