Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform

Claire A. Mitchell, Stefan Kalies, Tomás Cizmár, Alexander Heisterkamp, Lesley Torrance, Alison G. Roberts, Frank J. Gunn-Moore, Kishan Dholakia

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

A tightly-focused ultrashort pulsed laser beam incident upon a cell membrane has previously been shown to transiently increase cell membrane permeability while maintaining the viability of the cell, a technique known as photoporation. This permeability can be used to aid the passage of membrane-impermeable biologically-relevant substances such as dyes, proteins and nucleic acids into the cell. Ultrashort-pulsed lasers have proven to be indispensable for photoporating mammalian cells but they have rarely been applied to plant cells due to their larger sizes and rigid and thick cell walls, which significantly hinders the intracellular delivery of exogenous substances. Here we demonstrate and quantify femtosecond optical injection of membrane impermeable dyes into intact BY-2 tobacco plant cells growing in culture, investigating both optical and biological parameters. Specifically, we show that the long axial extent of a propagation invariant (''diffraction-free'') Bessel beam, which relaxes the requirements for tight focusing on the cell membrane, outperforms a standard Gaussian photoporation beam, achieving up to 70% optoinjection efficiency. Studies on the osmotic effects of culture media show that a hypertonic extracellular medium was found to be necessary to reduce turgor pressure and facilitate molecular entry into the cells.

Original languageEnglish
Article numbere79235
JournalPloS one
Volume8
Issue number11
DOIs
Publication statusPublished - 2013 Nov 14

Fingerprint

Tobacco
Cell membranes
tobacco
Cells
Plant Cells
Pulsed lasers
Lasers
Coloring Agents
Cell Membrane
Cell Membrane Permeability
Membranes
Gaussian beams
lasers
cells
cell membranes
dyes
Cell Wall
Nucleic Acids
Laser beams
Culture Media

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

Cite this

Mitchell, C. A., Kalies, S., Cizmár, T., Heisterkamp, A., Torrance, L., Roberts, A. G., ... Dholakia, K. (2013). Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform. PloS one, 8(11), [e79235]. https://doi.org/10.1371/journal.pone.0079235
Mitchell, Claire A. ; Kalies, Stefan ; Cizmár, Tomás ; Heisterkamp, Alexander ; Torrance, Lesley ; Roberts, Alison G. ; Gunn-Moore, Frank J. ; Dholakia, Kishan. / Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform. In: PloS one. 2013 ; Vol. 8, No. 11.
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Mitchell, CA, Kalies, S, Cizmár, T, Heisterkamp, A, Torrance, L, Roberts, AG, Gunn-Moore, FJ & Dholakia, K 2013, 'Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform', PloS one, vol. 8, no. 11, e79235. https://doi.org/10.1371/journal.pone.0079235

Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform. / Mitchell, Claire A.; Kalies, Stefan; Cizmár, Tomás; Heisterkamp, Alexander; Torrance, Lesley; Roberts, Alison G.; Gunn-Moore, Frank J.; Dholakia, Kishan.

In: PloS one, Vol. 8, No. 11, e79235, 14.11.2013.

Research output: Contribution to journalArticle

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Mitchell CA, Kalies S, Cizmár T, Heisterkamp A, Torrance L, Roberts AG et al. Femtosecond optoinjection of intact tobacco BY-2 cells using a reconfigurable photoporation platform. PloS one. 2013 Nov 14;8(11). e79235. https://doi.org/10.1371/journal.pone.0079235