Using electroporation and lipid-mediated transfection of GFP-expressing plasmids to label embryonic avian cells for vital confocal and two-photon microscopy.
Fluorescent proteins have emerged as an ideal fluorescent marker for studying cell morphologies in vital systems. These proteins were first applied in whole organisms with established germ-line transformation protocols, but now it is possible to label cells with fluorescent proteins in other organisms. Here we present two ways to introduce GFP expressing plasmids into avian embryos for vital confocal and two-photon imaging. First, electroporation is a powerful approach to introduce GFP into the developing neural tube, offering several advantages over dye labeling. Second, we introduce a new lipid-based transfection system for introducing plasmid DNA directly to a small group of injected cells within live, whole embryos. These complementary approaches make it possible to transfect a wide-range of cell types in the avian embryo and the bright, stable, uniform expression of GFP offers great advantages for vital fluorescence imaging.
Dickinson
Mary E
ME
Biological Imaging Center, Beckman Institute, California Institute of Technology, Pasadena 91125, USA. maryd@gg.caltech.edu
Murray
Ben A
BA
Haynes
Sherry M
SM
Waters
Christopher W
CW
Longmuir
Kenneth J
KJ
D000818:Animals; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D002465:Cell Movement; D002642:Chick Embryo / Q000166:cytology* / Q000378:metabolism; D018274:Electroporation / Q000379:methods*; D049452:Green Fluorescent Proteins; D007091:Image Processing, Computer-Assisted; D008055:Lipids / Q000737:chemistry; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D008853:Microscopy / Q000379:methods*; D018613:Microscopy, Confocal; D009432:Neural Crest / Q000166:cytology; D017785:Photons; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D014162:Transfection / Q000379:methods*
D015820:Cadherins; D008055:Lipids; D008164:Luminescent Proteins; D011994:Recombinant Proteins; C413663:fluorescent protein 583; D049452:Green Fluorescent Proteins