Modeling and experimental studies have shown that pulsed electric fields of nanosecond duration and megavolt per meter amplitude affect subcellular structures but do not lead to the formation of large pores in the outer membrane. This “intracellular electromanipulation” requires the use of pulse generators which provide extremely high power but low energy pulses. In this study, we describe the concept of the required pulsed power sources, their design, operation, and the necessary diagnostics. Two types of pulse generators based on the Blumlein line principle have been developed and are described here. One system is designed to treat a large number of cells in cuvettes holding volumes from 0.1 to 0.8 ml. Pulses of up to 40 kV amplitude, with a duration of 10 ns and a rise time close to 1 ns can be applied to the cuvette. For an electrode gap of 1 mm this voltage corresponds to an average electric field of 40 MV/m. The second system allows for real time observation of individual cells under a microscope. It generates pulses of 10–300 ns duration with a rise time of 3.5 ns and voltage amplitudes up to 1 kV. Connected to a microreactor with an electrode gap of 100 µm, electric fields up to 10 MV/m are applied. Bioelectromagnetics 27:172–187, 2006. © 2005 Wiley-Liss, Inc.