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We have studied the in vitro transfection of a plasmid DNA with thelacZgene to HeLa-S3 cells and hemolysis in a red blood cell (RBC) suspension under pulsed ultrasound with duty cycles?of 10, 20 and 30% using a digital sonifier at a frequency of 20 kHz and an intensity of 6.2 W/cm2 on the surface of a horn tip. Cultured HeLa-S3 cells in suspension were exposed to pulsed ultrasound for an apparent exposure timet' from 0 to 60 s. HeLa-S3 viability decreased as a single exponential function of the total exposure timet=?t' with a common time constantt=3.8 s for three duty cycles. Transfection was evaluated by counting the number of ß-galactosidase(ß-Gal)-positive cells relative to the total number of cells. Pulsed ultrasound provided an enhanced transfer of the ß-Gal plasmid to HeLa-S3 cells, 3.4-fold as compared with that in the case of the control. The optimal transfection efficiencies were 0.75, 0.80 and 0.74% neart=twith?=10, 20 and 30%, respectively. The number ratio of ß-Gal-positive cells to the surviving cells after exposure increased witht' according to a modified logistic equation. The degree of hemolysis also increased exponentially witht' at a time constantt'=t0/?for the RBC suspension in physiological saline at a hematocrit concentration of 0.5% witht0=0.9 s. Thus the total exposure time for the optimal transfection efficiency wast, that is, nearly four times oft0. Hemolysis in the RBC suspension may be a useful model for determining optimal transfection by pulsed ultrasound of various duty cycles. [ABSTRACT FROM AUTHOR]