Eppendorf Multiporator - Electroporation - Manual (Page 5)

29
The electroporation functions of the Multiporator ® are fundamentally different to those of other commercially available
devices. The main differences are as follows:
– Application of extremely short pulses, in the range of 15 to 100 microseconds, versus pulses in the millisecond range.
– Electronic pulse regulation, which allows uniform, reproducible pulses regardless of the resistance properties of the
media used.
– Electroporation in a hypoosmolar buffer which is non-toxic and which is adapted to the cytosolic ion composition of
the cells.
The combination of these features guarantees high transfection yields without severe damage to the cells. This contrasts
sharply with observations frequently made during electroporation in the millisecond range.
Soft Pulses
During electroporation, the membrane of a cell is charged up to a voltage at which the cell membrane is (reversibly)
permeated. The pulse lengths (i.e. the time constant
τ ) used with the Multiporator ® for plant and animal cells are usually
between 15 µs and 100 µs. In this period, the membrane is permeated when the permeation voltage is exceeded. This
leads to a drastic increase in the permeability of the membrane, which can be considered as pore-like openings in the
membrane.
If the external voltage applied is up to one thousand times longer (as is the case with milliseconds), high electrical
currents flowing through the inside of the cells inflict severe damage upon the cells themselves. Irreversible damage can
be caused to the membrane functions and the genome, as well as irreversible changes in the ion composition inside of
the pulsed cells. This is the situation with many conventional electroporation devices.
When the Soft Pulse technology of the Multiporator ® is applied, however, the cell is charged up only to the point at which
a breakthrough of the membrane occurs. The exponentially decaying pulse prevents significant amounts of current from
flowing through the cells after the pores have formed.
In addition, the Soft Pulse is measured continuously by the Multiporator ® and re-regulated every 5 µs. This unique
electronic regulation enables extremely high reproducibility.
As a result, the Multiporator ® ensures extremely high transfection rates.
Multiporator ® buffer system
Pulse media with low electrical conductivity ensure that the current flow is markedly reduced during electroporation, thus
preventing any significant damage to cells. In addition, such media guarantee that the electrically induced "pores" are
much larger than those obtained from pulses in conductive solutions, such as phosphate-buffered saline solutions (1).
The Multiporator ® is specially designed for the use of such low-conductivity pulse media.
Shifts in the pH value
If the current flow lasts a long time, electrolysis of water takes place on the electrodes of the cuvette. When millisecond
pulses are used, the pH value directly at the electrodes changes drastically. Shifts of the pH value in the alkaline and in
the acidic range are non-physiological and damage the cells. In contrast, no significant changes in the pH values in and
around the electrodes are noted when the Multiporator
® is used (2).
2 Principle of the Multiporator®
2 Principle of the Multiporator
®
Multipor_Appli_E_poration_en.fm Seite 29 Montag, 30. Januar 2006 2:17 14