Lipofectamine Transfection Protocol

Lipofectamine 2000 (Invitrogen) was used positive control in transfection ofluciferase in the cells. 5 pL of Lipofectamine 2000 was used for 2 pg of DNA to form a DNA complex in Opti-MEM, according to the manufacturer s protocol. 

An assessment of the transfection efficiency of human skin fibroblasts, compared to transfection of hamster lung fibroblasts and Human HeLa cells, indicated that the protocols we used are not suitable to obtain stable transfected skin fibroblasts. We concluded this from the fact that no stable transfected colonies of human skin fibroblasts were obtained, regardless of being CPTl-defective or being from a patient with a disorder unrelated to P-oxidation. Only transient expression with the Lipofectamine method was successful. We found an efficiency of 0.0-0.2 % of the cells which survived transfection without selection. However, compared to the efficiencies of transfection in hamster lung fibroblasts (5-10% in a parallel experiment), this still was low. Therefore, for long-term apphcations it is clear that different procedures should be used than the ones we applied here. 

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