Efficient nonviral gene delivery into primary lymphocytes from rats and mice

Authors:
Goffinet C and Keppler OT
In:
Source: FASEB J
Publication Date: (2006)
Issue: 20(3): 500-2
Research Area:
Cancer Research/Cell Biology
Immunotherapy / Hematology
Cells used in publication:
T cell, mouse - BALB/c
Species: mouse
Tissue Origin: blood
T cell, mouse, stim
Species: mouse
Tissue Origin: blood
Platform:
Nucleofectorâ„¢ I/II/2b
Abstract
Efficient nonviral gene delivery into primary lymphocytes from rats and mice. Goffinet C, Keppler OT. A rapid method for efficient gene delivery into primary rodent lymphocytes would greatly facilitate the study of signaling and metabolic pathways in untransformed hematopoietic cells as well as the validation of gene expression and targeting strategies before the generation of knockout or knock-down animals. Here, we report that species-adapted nucleofection procedures combined with optimized cultivation conditions render proliferating primary T cells, B cells, and natural killer cells from widely used rat and mouse strains susceptible to high-level gene delivery. As a result, transgene expression levels were enhanced ~10- to 370-fold over established protocols. The effectiveness of the nucleofection approach for functional analyses was demonstrated by specific down-regulation of CD4 cell surface molecules by either transient expression of the endocytosis-inducing Nef protein from human immunodeficiency virus or by specific gene silencing mediated by small interfering RNA. In conclusion, this species-adapted procedure for nonviral gene delivery renders primary rodent lymphocytes accessible to rapid functional ex vivo studies, which until now have not been feasible. Furthermore, nucleofection may aid the advancement of therapeutic nonviral gene delivery approaches.