| The field strength (V/cm) of the electrical pulse used is an essential
factor in determining the survival rate, as well as the transfection rate,
of the cells used.
If the field strength of the pulse exceeds a specific threshhold (= critical
external field strength), reversible permeation occurs in the cell membrane.
This so-called permeation voltage is heavily dependent on the temperature
at which electroporation takes place. The diagrams in Fig. 1 show
the minimum values of the pulse voltage which have to be set in relation
to the cell diameter and the temperature at which the electroporation
is performed. The diameter of the cell is determined after the cells have
been incubated in electroporation buffer for 10–15 minutes (see adjustment
of electroporation buffer). In addition, the gap width of the cuvettes
must be taken into account when the minimum pulse voltage is determined.
If the gap width is doubled, the pulse voltage must also be doubled in
order to obtain the same field strength. A general rule when determining
the ideal field strength is that small cells require a higher field strength
in order to achieve membrane permeation. The pulse voltages in Fig.1
and the corresponding Table 1 are the minimum values at which
the membrane can be permeated.
However, depending on the cell type used, optimal transfection efficiency
is often only achieved at significantly higher voltages. To determine
the optimal pulse voltage, it is advisable to carry out a series of experiments
in which the minimum value, twice the value and then three times the value
shown in Table 1 are used for suspension cells, and up to five
times the value for adherent cells.
Cells which do not assume a rounded form in the electroporation buffer
(see Fig. 2) often require even higher pulse voltages before optimal
transfection can occur.
Please note that increasing the pulse voltage can increase the transfection
rate but, at the same time, can also increase the cell mortality rate.
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