New trends in tumor biology: transfection of a human hepatoblastoma cell line with green fluorescent protein.
OBJECTIVE
Enhanced green fluorescent protein (eGFP) is widely used as a marker in different biologic systems. After vector transfection, eGFP is expressed by eukaryotic cells and can be visualized using fluorescent microscopy. The aim of this study was to establish an eGFP-transfected human hepatoblastoma (HB) cell line as tool for further basic research studies.


METHODS
The HB cell line HUH6 was transfected with the pEGFP-N1 vector by liposomal transfection. Enhanced green fluorescent protein-positive cells were sorted out by fluorescence-activated cell sort and selected using G418 resistance. Expression of eGFP-messenger RNA was assessed by single-cell reverse transcriptase polymerase chain reaction after laser microdissection. Original and transfected cells were compared biologically and cytomorphologically.


RESULTS
Vector transfection produced up to 15% eGFP-positive cells. After fluorescence-activated cell sort and G418 selection, a pure cell line was established with 100% eGFP-positive cells. Reverse transcriptase polymerase chain reaction revealed constant expression of eGFP-messenger RNA in fluorescending cells. Analysis of cell characteristics revealed no differences between transfected and original cells.


CONCLUSIONS
For the first time, the authors established an eGFP-transfected HB cell line. This cell line can serve as a promising tool for further studies investigating HB in vitro and in vivo. Our model might also be a basis for similar work on other pediatric solid tumors.
Warmann
Steven W
SW
Department of Pediatric Surgery, University of Tübingen, 72076 Tübingen, Germany. steven.warmann@med.uni-tuebingen.de <steven.warmann@med.uni-tuebingen.de>
Fuchs
Jörg
J
Seitz
Guido
G
Ruck
Peter
P
Treuner
Claudia
C
Mahrt
Jens
J
Müller
Gerhard A
GA
Wessels
Johannes T
JT
D005434:Flow Cytometry; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics*; D018197:Hepatoblastoma / Q000235:genetics*; D006801:Humans; D008081:Liposomes; D008113:Liver Neoplasms / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D014162:Transfection; D014407:Tumor Cells, Cultured
D008081:Liposomes; D012333:RNA, Messenger; D049452:Green Fluorescent Proteins