Testing for electrotransfection parameters by use of the fluorescent dye lucifer yellow CH.
A nonradioactive and rapid method to systematically optimize conditions for electrotransfection is described here for a critical parameter, the initial voltage. This technique utilizes the electric field-dependent transfer of the fluorescent compound Lucifer Yellow CH into cells. Dye uptake can be followed and quantified by fluorescence microscopy for individual cells or in sum by fluorescence spectroscopy. Electrotransfection conditions for maximal dye and DNA uptake correspond with each other. Cotransfection of Lucifer Yellow CH and DNA coding for the indicator gene chloramphenicol acetyltransferase followed by fluorescence-activated cell sorting demonstrates that the cell population that takes up the fluorescent compound also expresses the indicator gene.
Sczakiel
G
G
Institut für Virusforschung, Deutsches Krebsforschungszentrum, Heidelberg, Federal Republic of Germany.
Döffinger
R
R
Pawlita
M
M
D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis / Q000235:genetics; D004247:DNA / Q000032:analysis / Q000235:genetics; D004574:Electromagnetic Fields; D006801:Humans; D007546:Isoquinolines; D008856:Microscopy, Fluorescence; D013050:Spectrometry, Fluorescence; D014162:Transfection; D014407:Tumor Cells, Cultured
D007546:Isoquinolines; D004247:DNA; C017475:lucifer yellow; D015500:Chloramphenicol O-Acetyltransferase