( James R. Wong Ph.D. and...)Transient Gene Expression Following Single and Double Exponential Decay Electroporation Pulses

James R. Wong, Ph.D. and Wei-Yu Lin, Bio-Rad Laboratories

Summary
A double pulse format for electroporation of mammalian tissue culture cells gave lower transient expression than a single exponential decay pulse. Moreover, separate administration of the two components of the double pulse revealed that transformation was due almost exclusively to the first, high voltage pulse and not to the second, long duration waveform.

Introduction
The Twin-pulse wave delivery is a novel electroporation waveform consisting of a short duration, exponential decay pulse followed by a second capacitor discharge yielding a long duration pulse. It is claimed that the first high-voltage pulse creates pores, or openings in the cell walls due to the high voltage delivered... (and) the second low-voltage pulse delivers energy to those cells shocked by the first pulse, therefore favoring entry of exogenous molecules into the cells by means of creating an active electrophoresis field.¹ While the transient formation of pores in the plasma membrane following delivery of appropriate electrical fields has been documented via electron microscopy,² the mechanism of entry and egress of materials through these pores is not known, and no evidence has been provided for the postulated Twin-pulse description.

De Chasseval and de Villartay described the use of double electroporation pulse format to introduce pairs of plasmids into human lymphoid cells.³ Their principal finding was that cotransfection of a plasmid encoding the SV40 large T antigen with a reporter plasmid bearing the SV40 origin of replication increased transient lucifer
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