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Cited by 117Microbial Inactivation MethodsRNA Interference and Gene DeliveryPlasma Applications and Diagnostics

The effects of electroporation buffer composition on cell viability and electro-transfection efficiency

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Sherba, Joseph J., Hogquist, Stephen, Lin, Hao, Shan, Jerry W., Shreiber, David I., Zahn, Jeffrey D.·2020·Scientific Reports
DOI 10.1038/s41598-020-59790-xPMC PMC7033148
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Abstract

Electroporation is an electro-physical, non-viral approach to perform DNA, RNA, and protein transfections of cells. Upon application of an electric field, the cell membrane is compromised, allowing the delivery of exogenous materials into cells. Cell viability and electro-transfection efficiency (eTE) are dependent on various experimental factors, including pulse waveform, vector concentration, cell type/density, and electroporation buffer properties. In this work, the effects of buffer composition on cell viability and eTE were systematically explored for plasmid DNA encoding green fluorescent protein following electroporation of 3T3 fibroblasts. A HEPES-based buffer was used in conjunction with various salts and sugars to modulate conductivity and osmolality, respectively. Pulse applications were chosen to maintain constant applied electrical energy (J) or total charge flux (C/m 2 ). The energy of the pulse application primarily dictated cell viability, with Mg 2+ -based buffers expanding the reversible electroporation range. The enhancement of viability with Mg 2+ -based buffers led to the hypothesis that this enhancement is due to ATPase activation via re-establishing ionic homeostasis. We show preliminary evidence for this mechanism by demonstrating that the enhanced viability is eliminated by introducing lidocaine, an ATPase inhibitor. However, Mg 2+ also hinders eTE compared to K + -based buffers. Collectively, the results demonstrate that the rational selection of pulsing conditions and buffer compositions are critical for the design of electroporation protocols to maximize viability and eTE.

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  • Disadvantages of non-viral vector and DNA transfection systems include process complexity, need for significant method optimization to overcome low efficiency of CAR expression, need for cell sorting post-engineering, and significant cell loss during gene delivery.

    “Disadvantage of these systems include process complexity, need for significant method optimization to overcome low efficiency of CAR expression, need for cell sorting post-engineering (Table S1) and significant cell loss during gene delivery (106).”

    From Scalable Manufacturing of CAR T Cells for Cancer Immunotherapy (2021) · cites this paper as [106]

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References 40 · Cited by 117

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  1. [1]Carlo DD Lee LP Dynamic Single-Cell Analysis for Quantitative Biology Analytical Chemistry 2006 78 7918 7925 10.1021/ac069490p 17186633 · DOI
  2. [2]Wang D Bodovitz S Single cell analysis: The new frontier in “omics” Trends in Biotechnology 2010 28 281 290 10.1016/j.tibtech.2010.03.002 20434785 PMC2876223 · DOI
  3. [3]Zheng M Continuous-flow, electrically-triggered, single cell-level electroporation Technology 2017 5 31 41 10.1142/S2339547817500017 · DOI
  4. [4]Ginn SL Gene therapy clinical trials worldwide to 2012 - an update Journal of Gene Medicine 2013 15 65 77 10.1002/jgm.2698 23355455 · DOI
  5. [5]Gallego-Perez D Topical tissue nano-transfection mediates non-viral stroma reprogramming and rescue Nature Nanotechnology 2017 12 974 979 10.1038/nnano.2017.134 PMC5814120 28785092 · DOI
  6. [6]Kim TK Eberwine JH Mammalian cell transfection: The present and the future Analytical and Bioanalytical Chemistry 2010 397 3173 3178 10.1007/s00216-010-3821-6 20549496 PMC2911531 · DOI · Paper page
  7. [7]Roth TL Reprogramming human T cell function and specificity with non-viral genome targeting Nature 2018 559 405 409 10.1038/s41586-018-0326-5 29995861 PMC6239417 · DOI · Paper page
  8. [8]Bolhassani, A., Khavari, A. & Orafa, Z. Electroporation – Advantages and Drawbacks for Delivery of Drug, Gene and Vaccine. In Application of Nanotechnology in Drug Delivery. A.D. Sezer (Ed.). (2014), InTech, Rijeka, Croatia - EU.
  9. [9]Kotnik T Membrane Electroporation and Electropermeabilization: Mechanisms and Models Annual Review of Biophysics 2019 48 63 91 10.1146/annurev-biophys-052118-115451 30786231 · DOI · Paper page
  10. [10]Djuzenova CS Effect of medium conductivity and composition on the uptake of propidium iodide into electropermeabilized myeloma cells. Biochim. Biophys. Acta Biomembranes 1996 1284 143 152 10.1016/S0005-2736(96)00119-8 8914578 · DOI
  11. [11]Yao S Improvement of electroporation to deliver plasmid DNA into dental follicle cells Biotechnology Journal 2009 4 1488 1496 10.1002/biot.200900039 19830717 PMC2824253 · DOI
  12. [12]Pucihar G Equivalent Pulse Parameters for Electroporation IEEE Transactions on Biomedical Engineering 2011 58 3279 3288 10.1109/TBME.2011.2167232 21900067 · DOI
  13. [13]Potter H Heller R Transfection by Electroporation Curr Protoc Mol Biol. 2003 9 1 11 20890905
  14. [14]Ding X High-throughput Nuclear Delivery and Rapid Expression of DNA via Mechanical and Electrical Cell-Membrane Disruption Nature Biomedical Engineering 2017 1 0039 10.1038/s41551-017-0039 PMC5602535 28932622 · DOI
  15. [15]Li LH Highly efficient, large volume flow electroporation Technology in Cancer Research and Treatment 2002 1 341 349 10.1177/153303460200100504 12625759 · DOI
  16. [16]Demiryurek Y Transport, resealing, and re-poration dynamics of two-pulse electroporation-mediated molecular delivery Biochimica et Biophysica Acta (BBA) – Biomembranes 2015 1848 1706 1714 10.1016/j.bbamem.2015.04.007 25911207 · DOI
  17. [17]Li J Lin H Numerical simulation of molecular uptake via electroporation Bioelectrochemistry 2011 82 10 21 10.1016/j.bioelechem.2011.04.006 21621484 · DOI
  18. [18]Li J The effect of extracellular conductivity on electroporation-mediated molecular delivery Biochimica et Biophysica Acta - Biomembranes 2013 1828 461 470 10.1016/j.bbamem.2012.08.014 22954677 · DOI
  19. [19]Sadik MM Quantification of propidium iodide delivery using millisecond electric pulses: Experiments Biochimica et Biophysica Acta – Biomembranes 2013 1828 1322 1328 10.1016/j.bbamem.2013.01.002 23313458 · DOI
  20. [20]Zheng M Hydrodynamically controlled cell rotation in an electroporation microchip to circumferentially deliver molecules into single cells Microfluidics and Nanofluidics 2016 20 1 12 10.1007/s10404-015-1676-z · DOI
  21. [21]Sadik MM Scaling relationship and optimization of double-pulse electroporation Biophysical Journal 2014 106 801 812 10.1016/j.bpj.2013.12.045 24559983 PMC3944924 · DOI
  22. [22]Haberl S Miklavčič D Pavlin M Effect of Mg ions on efficiency of gene electrotransfer and on cell electropermeabilization Bioelectrochemistry 2010 79 265 271 10.1016/j.bioelechem.2010.04.001 20580903 · DOI
  23. [23]Haberl S Effect of different parameters used for in vitro gene electrotransfer on gene expression efficiency, cell viability and visualization of plasmid DNA at the membrane level The Journal of Gene Medicine 2013 15 169 181 10.1002/jgm.2706 23564663 · DOI
  24. [25]Wang H Griffiths MW Mg2+-free buffer elevates transformation efficiency of Vibrio parahaemolyticus by electroporation Letters in Applied Microbiology 2009 48 349 354 10.1111/j.1472-765X.2008.02531.x 19207855 · DOI
  25. [26]Wu M Yuan F Membrane Binding of Plasmid DNA and Endocytic Pathways Are Involved in Electrotransfection of Mammalian Cells PLOS ONE 2011 6 1 9 PMC3113837 21695134
  26. [27]Schmitz C Dual-Function Ion Channel/Protein Kinases: Novel Components of Vertebrate Magnesium Regulatory Mechanisms Pediatric Research 2004 55 734 737 10.1203/01.PDR.0000117848.37520.A2 14764909 · DOI
  27. [28]Ku WY The zinc ion in the HNH motif of the endonuclease domain of colicin E7 is not required for DNA binding but is essential for DNA hydrolysis Nucleic Acids Research 2002 30 1670 1678 10.1093/nar/30.7.1670 11917029 PMC101835 · DOI
  28. [29]Delgado-Cañedo A Optimization of an electroporation protocol using the K562 cell line as a model: Role of cell cycle phase and cytoplasmic DNAses Cytotechnology 2006 51 141 148 10.1007/s10616-006-9028-1 19002884 PMC3449805 · DOI
  29. [30]Reinhart RA Clinical correlates of the molecular and cellular actions of magnesium on the cardiovascular system American Heart Journal 1991 121 1513 1521 10.1016/0002-8703(91)90160-J 2017983 · DOI
  30. [31]Rols M Control by ATP and ADP of voltage-induced mammalian-cell-membrane permeabilization, gene transfer and resulting expression European Journal of Biochemistry 1998 254 2 382 388 10.1046/j.1432-1327.1998.2540382.x 9660195 · DOI
  31. [32]Pilotelle-Bunner A Mechanism of Mg2+binding in the Na+, K+-ATPase Biophysical Journal 2009 96 3753 3761 10.1016/j.bpj.2009.01.042 19413981 PMC2711396 · DOI
  32. [33]Yatime L P-type ATPases as drug targets: Tools for medicine and science Biochimica et Biophysica Acta - Bioenergetics 2009 1787 207 220 10.1016/j.bbabio.2008.12.019 19388138 · DOI
  33. [34]Grys M Madeja Z Korohoda W Decreasing the thresholds for electroporation by sensitizing cells with local cationic anesthetics and substances that decrease the surface negative electric charge Cellular and Molecular Biology Letters 2014 19 65 76 10.2478/s11658-013-0114-z 24415057 PMC6275628 · DOI
  34. [35]Kutchai H Geddis LM Farley RA Effects of Local Anaesthetics on the Activity of the Na, K-ATPase of Canine Renal Medulla Pharmacological Research 2000 41 1 7 10.1006/phrs.1999.0547 10600263 · DOI
  35. [36]Rosazza C Gene Electrotransfer: A Mechanistic Perspective Current Gene Therapy 2016 16 98 129 10.2174/1566523216666160331130040 27029943 PMC5412002 · DOI
  36. [37]Mao M Involvement of a Rac1-Dependent Macropinocytosis Pathway in Plasmid DNA Delivery by Electrotransfection Molecular Therapy 2017 25 803 815 10.1016/j.ymthe.2016.12.009 28129959 PMC5363188 · DOI
  37. [38]Rols MP Temperature effects on electrotransfetion of mammalian cells Nucleic Acids Research 1994 22 540 10.1093/nar/22.3.540 8127697 PMC523619 · DOI

Authors
Sherba, J. J., Hogquist, S., Lin, H., Shan, J. W., Shreiber, D. I., & Zahn, J. D.
Affiliations
Rutgers, The State University of New Jersey (US)
Corresponding author
Zahn, Jeffrey D.
ORCID
Venue
Scientific Reports
Publisher
Springer Science and Business Media LLC
Published
2020-02-20
Volume
10
Issue
1
Pages
3053-3053
ISSN
2045-2322 (ISSN-L)
DOI
10.1038/s41598-020-59790-x
PMC
PMC7033148
OpenAlex
W3007613875
Cited by
117
References
40
Citations by year
2026: 16 · 2025: 16 · 2024: 15 · 2023: 18 · 2022: 26 · 2021: 19 · 2020: 7
FWCI
9.06
Type
journal article
Language
en
Open access
Licence
Locations
  • Scientific Reports (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Rutgers University Community Repository (Rutgers University) (submittedVersion, cc-by)
Funders
National Science Foundation (DBI IDBR 135391, CBET 0967598)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biotechnology › Microbial Inactivation Methods
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Plasma Applications and Diagnostics
Keywords
electroporation, cell viability, plasmid transfection, 3T3 fibroblasts, HEPES buffer, magnesium ions, potassium ions, osmolality, conductivity, pulse energy, ATPase activation, ion homeostasis, lidocaine, ATPase inhibitors, green fluorescent protein
MeSH terms
Adenosine Triphosphatases, Animals, Buffers, Cell Survival, Electricity, Magnesium, Transfection, Electroporation, NIH 3T3 Cells, Mice

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