Human cultured (stimulated) T cells
When stimulated by CD3 and CD28 antibodies, T cells grow much larger than fresh PBMC cells. Therefore the voltages used for cultured T cell electroporation is much lower than fresh PBMCs.
We have noticed that T cells cultured in different labs require different voltages for electroporation. The variation may be caused by different brands of culture mediums or different CD3/CD28 antibody and IL-2 combination.
To achieve a good transfection efficiency for cultured T cells while maintaining a good cell proliferative potential, it is imperative that the electroporation voltages be tested out.
A special notice for using transposon system with T cells: we noticed that a sleeping beauty transposon system works at a much lower efficiency in stimulated T cells than in unstimulated PBMCs. If you are working with a transposon system such as sleeping beauty or piggybac and having trouble with expression in cultured T cells, try your expression system in fresh PBMCs and it might work much better.
Transfection efficiency with GFP plasmid after 5 days is over 90%
T cell, cultured or stimulated
in 20ul tubes
Cell Number：1-2*10^6 cells
Plasmid：2ug
E buffer：Celetrix buffer
Suggested parameter：Cell Line Mode，480V-540V
Culture medium: antiCD3/antiCD28/IL2 (antibiotics free)
T cell, cultured or stimulated
in 100ul tubes
Cell Number：5-10*10^6 cells
Plasmid：10ug
E buffer：Celetrix buffer
Suggested parameter：Cell Line Mode，480V-540V
Culture medium: antiCD3/antiCD28/IL2 (antibiotics free)