Topic
Optical transfection
About: Optical transfection is a research topic. Over the lifetime, 40 publications have been published within this topic receiving 2200 citations.
Papers
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TL;DR: A variety of mammalian cells can be directly transfected with DNA without perturbing their structure by first creating a tiny, localized perforation in the membrane using ultrashort (femtosecond), high-intensity, near-infrared laser pulses.
Abstract: The challenge for successful delivery of foreign DNA into cells in vitro, a key technique in cell and molecular biology with important biomedical implications, is to improve transfection efficiency while leaving the cell's architecture intact. Here we show that a variety of mammalian cells can be directly transfected with DNA without perturbing their structure by first creating a tiny, localized perforation in the membrane using ultrashort (femtosecond), high-intensity, near-infrared laser pulses. Not only does this superior optical technique give high transfection efficiency and cell survival, but it also allows simultaneous evaluation of the integration and expression of the introduced gene.
618 citations
TL;DR: Contrary to recent literature, in which 100% efficiency is claimed, this measure of efficiency accounts for all irradiated cells, including those lost as a result of laser treatment, thereby providing a true biological measure of the technique.
Abstract: Photoporation is a rapidly expanding technique for the introduction of macromolecules into single cells. However, there remains no study into the true efficiency of this procedure. Here, we present a detailed analysis of transfection efficiency and cell viability for femtosecond optical transfection using a titanium sapphire laser at 800 nm. Photoporation of 4000 Chinese Hamster ovary cells was performed, representing the largest optical transfection study reported to date. We have investigated a range of laser fluences at the cell membrane and, at 1.2 microJ/cm(2), have found an average transfection efficiency of 50 +/- 10%. Contrary to recent literature, in which 100% efficiency is claimed, our measure of efficiency accounts for all irradiated cells, including those lost as a result of laser treatment, thereby providing a true biological measure of the technique.
213 citations
TL;DR: In this paper, a frequency-multiplied Nd:YAG laser, 355 m wavelength, 5 ns pulse duration, punches a self-healing hole of submicrometer aperture in cell membrane under selected irradiation conditions.
Abstract: A new technique is presented to incorporate exogeneous gene materials (DNA) into cells with a microbeam irradiation from an uv pulsed laser. A frequency-multiplied Nd:YAG laser, 355 m wavelength, 5 ns pulse duration, punches a self-healing hole of submicrometer aperture in cell membrane under selected irradiation conditions. It takes a fraction of a second for the aperture to close, long enough to allow the foreign DNA, contained in the medium, to slip into the cell. The method offers a clear advantage over existing methods: increases the success rate of DNA transfection as well as the efficiency of cell modification by orders of magnitude.
196 citations
TL;DR: This discussion explores a procedure called optical injection, where a laser field transiently increases the membrane permeability to allow species to be internalized and provides a forecast of future applications of this rapidly developing and exciting technology.
Abstract: The plasma membrane of a eukaryotic cell is impermeable to most hydrophilic substances, yet the insertion of these materials into cells is an extremely important and universal requirement for the cell biologist. To address this need, many transfection techniques have been developed including viral, lipoplex, polyplex, capillary microinjection, gene gun and electroporation. The current discussion explores a procedure called optical injection, where a laser field transiently increases the membrane permeability to allow species to be internalized. If the internalized substance is a nucleic acid, such as DNA, RNA or small interfering RNA (siRNA), then the process is called optical transfection. This contactless, aseptic, single cell transfection method provides a key nanosurgical tool to the microscopist—the intracellular delivery of reagents and single nanoscopic objects. The experimental possibilities enabled by this technology are only beginning to be realized. A review of optical transfection is presented, along with a forecast of future applications of this rapidly developing and exciting technology.
178 citations
TL;DR: A Bessel beam is used that obviates the need to locate precisely the cell membrane, permitting two-photon excitation along a line leading to cell transfection, and demonstrates celltransfection beyond obstacles due to the self-healing nature of the Besselbeam.
Abstract: The ability to permeate selectively the cell membrane and introduce therapeutic agents is a key goal in cell biology. Optical transfection is a powerful methodology but requires exact focusing due to the required two-photon power density. The authors use a Bessel beam that obviates the need to locate precisely the cell membrane, permitting two-photon excitation along a line leading to cell transfection. Assuming a minimum efficiency of 20%, the Bessel beam offers transfection at axial distances 20 times greater than that of its Gaussian equivalent. Furthermore, the authors demonstrate cell transfection beyond obstacles due to the self-healing nature of the Bessel beam.
148 citations