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[h=2]Introduction[/thanks]Transfection is a process whereby foreign genetic material is introduced into eukaryotic cells. To this end, several transfection techniques have been developed. However, irrespective of the method used, the transfection efficiency is not 100%.
Although this issue can be overcome to some extent by stable transfection, variability in the transfection efficiency is unavoidable in the case of transient transfection. This means that to compare the results of different experiments, the transfection efficiency must be determined every time transient transfection is performed.
One easy and widely used technique to determine the transfection efficiency involves the use of the GFP gene. By forming a fusion protein between the protein of interest and GFP, the expression of the protein of interest can be determined by measuring the presence of GFP protein.
[h=2]Quantification of GFP Transfection Efficiency[/thanks]The Luna-FL dual fluorescence cell counter is an advanced instrument (Figure 1-A), fitted with an optical module for fluorescence signal and bright field signal detection.
Fluorescence channel 1 (FL1) is sufficient for stimulating GFP and receiving the light emitted from GFP. It also picks up signals emitted from Yellow Fluorescence Protein (YFP), although the excitation efficiency is less than with GFP.
In order to determine the GFP transfection efficiency with the Luna-FL fluorescence cell counter, 10
Although this issue can be overcome to some extent by stable transfection, variability in the transfection efficiency is unavoidable in the case of transient transfection. This means that to compare the results of different experiments, the transfection efficiency must be determined every time transient transfection is performed.
One easy and widely used technique to determine the transfection efficiency involves the use of the GFP gene. By forming a fusion protein between the protein of interest and GFP, the expression of the protein of interest can be determined by measuring the presence of GFP protein.
[h=2]Quantification of GFP Transfection Efficiency[/thanks]The Luna-FL dual fluorescence cell counter is an advanced instrument (Figure 1-A), fitted with an optical module for fluorescence signal and bright field signal detection.
Figure 1-A. Luna-FL Dual Fluorescence Cell Counter
Figure 1-B. Schematic of optical module of Luna-FL
Figure 1-B shows the schematic layout of the optical module of Luna-FL. Table 1 lists the optical specification of Luna-FL fluorescence cell counter.
Table 1. Optical specification of Luna-FL
| Excitation | 470/40nm |
| Dichroic (cutoff) | 495nm |
| Emission (FL1) | 530/50nm |
| Emission (FL2) | 600LP |
In order to determine the GFP transfection efficiency with the Luna-FL fluorescence cell counter, 10