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Interrelation between HeLa-S3 cell transfection and hemolysis in red blood cell suspension using pulsed ultrasound of various duty cycles

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Abstract

We have studied the in vitro transfection of a plasmid DNA with the lacZ gene to HeLa-S3 cells and hemolysis in a red blood cell (RBC) suspension under pulsed ultrasound with duty cycles γ 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 time t′ from 0 to 60 s. HeLa-S3 viability decreased as a single exponential function of the total exposure time t=γt′ with a common time constant τ=3.8 s for three duty cycles. Transfection was evaluated by counting the number of β-galactosidase(β-Gal)-positive cells relative to the total number of cells. Pulsed ultrasound provided an enhanced transfer of the β-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% near t=τ with γ=10, 20 and 30%, respectively. The number ratio of β-Gal-positive cells to the surviving cells after exposure increased with t′ according to a modified logistic equation. The degree of hemolysis also increased exponentially with t′ at a time constant τ′=τ0/γ for the RBC suspension in physiological saline at a hematocrit concentration of 0.5% with τ0=0.9 s. Thus the total exposure time for the optimal transfection efficiency was τ, that is, nearly four times of τ0. Hemolysis in the RBC suspension may be a useful model for determining optimal transfection by pulsed ultrasound of various duty cycles.

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Acknowledgements

The authors would like to thank Dr Toshihiko Saheki for instruction on plasmid vector preparation. We also thank Dr Yoshiharu Toyama for helpful discussions.

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Authors and Affiliations

  1. Department of Biological and Chemical Engineering, Faculty of Engineering, Gunma University, Kiryu, Gunma 376-8515, Japan

    Y. Liu, H. Uno, H. Takatsuki, M. Hirano & A. Sakanishi

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  1. Y. Liu
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  2. H. Uno
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  3. H. Takatsuki
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  4. M. Hirano
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  5. A. Sakanishi
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Correspondence to A. Sakanishi.

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Liu, Y., Uno, H., Takatsuki, H. et al. Interrelation between HeLa-S3 cell transfection and hemolysis in red blood cell suspension using pulsed ultrasound of various duty cycles. Eur Biophys J 34, 163–169 (2005). https://doi.org/10.1007/s00249-004-0439-7

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  • DOI: https://doi.org/10.1007/s00249-004-0439-7

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