TY - JOUR
T1 - High-efficiency nonviral transfection of primary chondrocytes.
AU - Welter, Jean F.
AU - Solchaga, Luis A.
AU - Stewart, Matthew C.
PY - 2004
Y1 - 2004
N2 - The introduction of foreign DNA into mammalian cells is an essential investigative tool in molecular biology. Nonviral approaches to transfection offer the advantage of relatively simple vector design, production, and purification and, for tissue engineering applications, avoid many of the potential risks associated with virus-mediated transfection methods. Unfortunately, primary cells, and in particular chondrocytes, are notoriously refractory to conventional transfection approaches, and optimized transfection efficiencies in these cells are extremely low (1-1.5%). In this chapter, we present three protocols that have proved useful in transfecting primary chondrocytes at high efficiency (~70%). The first uses radiofrequency electroporation, a transfection method that frequently works extremely well in cell types that are difficult to transfect. It should be noted that electroporation is not limited to DNA but that essentially any molecule can be introduced into the cell using this approach. In addition to the primary protocol, we present two additional reliable, albeit less efficient backup protocols, the first using exponential decay electroporation and the second FuGENE 6 transfection.
AB - The introduction of foreign DNA into mammalian cells is an essential investigative tool in molecular biology. Nonviral approaches to transfection offer the advantage of relatively simple vector design, production, and purification and, for tissue engineering applications, avoid many of the potential risks associated with virus-mediated transfection methods. Unfortunately, primary cells, and in particular chondrocytes, are notoriously refractory to conventional transfection approaches, and optimized transfection efficiencies in these cells are extremely low (1-1.5%). In this chapter, we present three protocols that have proved useful in transfecting primary chondrocytes at high efficiency (~70%). The first uses radiofrequency electroporation, a transfection method that frequently works extremely well in cell types that are difficult to transfect. It should be noted that electroporation is not limited to DNA but that essentially any molecule can be introduced into the cell using this approach. In addition to the primary protocol, we present two additional reliable, albeit less efficient backup protocols, the first using exponential decay electroporation and the second FuGENE 6 transfection.
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U2 - 10.1385/1-59259-810-2:129
DO - 10.1385/1-59259-810-2:129
M3 - Article
C2 - 15280593
AN - SCOPUS:16644386874
SN - 1543-1894
VL - 100
SP - 129
EP - 146
JO - Methods in molecular medicine
JF - Methods in molecular medicine
ER -