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Reversible permeabilization using high‐intensity femtosecond laser pulses: Applications to biopreservation
Author(s) -
Kohli Vikram,
Acker Jason P.,
Elezzabi Abdulhakem Y.
Publication year - 2005
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.20689
Subject(s) - femtosecond , cryoprotectant , biopreservation , biophysics , laser , chemistry , biology , cryopreservation , microbiology and biotechnology , optics , embryo , bacteriocin , physics , antimicrobial , organic chemistry
Non‐invasive manipulation of live cells is important for cell‐based therapeutics. Herein we report on the uniqueness of using high‐intensity femtosecond laser pulses for reversibly permeabilizing mammalian cells for biopreservation applications. When mammalian cells were suspended in a impermeable hyperosmotic cryoprotectant sucrose solution, femtosecond laser pulses were used to transiently permeabilize cells for cytoplasmic solute uptake. The kinetics of cells exposed to 0.2, 0.3, 0.4, and 0.5 M sucrose, following permeabilization, were measured using video microscopy, and post‐permeabilization survival was determined by a dual fluorescence membrane integrity assay. Using appropriate laser parameters, we observed the highest cell survival for 0.2 M sucrose solution (>90%), with a progressive decline in cell survival towards higher concentrations. Using diffusion equations describing the transport of solutes, the intracellular osmolarity at the inner surface of the membrane ( x = 10 nm) and to a diffusive length of x = 10 μm was estimated, and a high loading efficiency (>98% for x = 10 nm and >70% for x = 10 μm) was calculated for cells suspended in 0.2 M sucrose. This is the first report of using femtosecond laser pulses for permeabilizing cells in the presence of cryoprotectants for biopreservation applications. © 2005 Wiley Periodicals, Inc.