A novel method of DNA transfection by laser microbeam cell surgery

Tsukakoshi, Motowo; Kurata, Shigeaki; Nomiya, Yoshio; Ikawa, Yoji; Kasuya, Tadashi

doi:10.1007/bf00697702

Cited by 162 publications

(114 citation statements)

References 11 publications

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“…It has been previously shown that the transfection efficiency decreases significantly when transfected with a plasmid concentration less than 10 mg/ml [3,15]. However we show that, in the presence of Nupherin-neuron, we could lower the plasmid concentration to 3.3 mg/ml without compromising the resulting transfection efficiency.…”

Section: Biophotonicscontrasting

confidence: 55%

“…[2], nanosecond lasers [3] and femtosecond lasers [4][5]. The physical mechanisms for transfection by each laser type differs.…”

Section: Biophotonicsmentioning

confidence: 99%

“…It was also observed that the addition of Nupherin-neuron did not affect the rate of cell proliferation. The corrected transfection efficiency was evaluated by the equation N cor ¼ ((E/D) 100)/2 tf , where t is the unit time and f is the frequency of division in unit time (In this case unit of time has been taken as 'days') [3]. Figure 6 shows a comparison between the transfection efficiencies of adherent and non-adherent CHO-K1 cells, corrected for the rate of cell proliferation.…”

Section: Transfection With Non-adherent Cho-k1 Cellsmentioning

confidence: 99%

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Enhancement and optimization of plasmid expression in femtosecond optical transfection

Praveen

Stevenson

Antkowiak

et al. 2011

Journal of Biophotonics

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Cell transfection using femtosecond lasers is gaining importance for its proven ability to achieve selective transfection in a sterile and relatively non‐invasive manner. However, the net efficiency of this technique is limited due to a number of factors that ultimately makes it difficult to be used as a viable and widely used technique. We report here a method to achieve significant enhancement in the efficiency of femtosecond optical transfection. The transfection procedure is modified by incorporating a suitable synthetic peptide containing nuclear localization and DNA binding sequences, assisting DNA import into the nucleus. We achieved a 3‐fold enhancement in the transfection efficiency for adherent Chinese Hamster Ovary (CHO‐K1) cells with this modified protocol. Further, in the presence of this biochemical reagent, we were able to reduce the required plasmid concentration by ∼70% without compromising the transfection efficiency. Also, we report for the first time the successful photo‐transfection of recently trypsinised cells with significantly high transfection efficiency when transfected with modified plasmid. This paves the way for the development of high throughput microfluidic optical transfection devices. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

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Section: Biophotonicscontrasting

confidence: 55%

“…[2], nanosecond lasers [3] and femtosecond lasers [4][5]. The physical mechanisms for transfection by each laser type differs.…”

Section: Biophotonicsmentioning

confidence: 99%

Section: Transfection With Non-adherent Cho-k1 Cellsmentioning

confidence: 99%

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Enhancement and optimization of plasmid expression in femtosecond optical transfection

Praveen

Stevenson

Antkowiak

et al. 2011

Journal of Biophotonics

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“…Their operation is usually based on the creation of a shock wave [11,12] which limits the selectivity of the method, although single cell nanosecond transfection experiments have been reported [13]. The lack of selectivity is compensated with a higher throughput and injection efficiency in some cases [14].…”

Section: Biophotonicsmentioning

confidence: 99%

Application of dynamic diffractive optics for enhanced femtosecond laser based cell transfection

Antkowiak

Torres-Mapa

Gunn‐Moore

et al. 2010

Journal of Biophotonics

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We demonstrate the advantages of a dynamic diffractive optical element, namely a spatial light modulator (SLM) for the controlled and enhanced optoinjection and phototransfection of mammalian cells with a femtosecond light source. The SLM provides full control over the lateral and axial positioning of the beam with sub‐micron precision. Fast beam translation enables time‐sequenced irradiation, which is shown to enhance the optoinjection efficiency and alleviate the problem of exact beam positioning on the cell membrane. We show that irradiation in three axial positions doubles the number of viably optoinjected cells when compared with a single dose. The presented system also enables untargeted raster scan irradiation which provides a higher throughput transfection of adherent cells at the rate of 1 cell per second. Additionally, fluorescent imaging is used to demonstrate cell selective two‐step gene therapy. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

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“…Building on the knowledge from medical laser uses and robust high-throughput semiconductor manufacturing tools, a novel laser-mediated cell manipulation technology has been developed. The resulting laser-enabled analysis and processing (LEAP) technology can implement cell imaging and multiple forms of laser-mediated cell manipulations including cell purification, optoinjection (i.e., laser-mediated cell transfection) (18,19), and chromophore-assisted laser-inactivation (e.g., direct knockout of protein function) (20) in an automated and highthroughput manner, thereby overcoming certain limitations of previous approaches to these methods (12,(21)(22)(23).…”

mentioning

confidence: 99%

High‐throughput laser‐mediated in situ cell purification with high purity and yield

et al. 2004

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Background: Technologies for purification of living cells have significantly advanced basic and applied research in many settings. Nevertheless, certain challenges remain, including the robust and efficient purification (e.g., high purity, yield, and sterility) of adherent and/or fragile cells and small cell samples, efficient cell cloning, and safe purification of biohazardous cells. In addition, existing purification methods are generally open loop and exhibit an inverse relation between cell purity and yield. Methods: An automated closed-loop (i.e., employing feedback control) cell purification technology was developed by building upon medical laser applications and laser-based semiconductor manufacturing equipment. Laser-enabled analysis and processing has combined highthroughput in situ cell imaging with laser-mediated cell manipulation via large field-of-view optics and galvanometer steering. Laser parameters were determined for cell purification using three mechanisms (photothermal, photochemical, and photomechanical), followed by demonstration of system performance and utility. Results: Photothermal purification required approximately 10 8 W/cm 2 at 523 nm in the presence of Allura Red, resulting in immediate protein coagulation and cell necrosis. Photochemical purification required approximately 10 9 W/cm 2 at 355 nm, resulting in apoptosis induction over 4 to 24 h. Photomechanical purification

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A novel method of DNA transfection by laser microbeam cell surgery

Cited by 162 publications

References 11 publications

Enhancement and optimization of plasmid expression in femtosecond optical transfection

Enhancement and optimization of plasmid expression in femtosecond optical transfection

Application of dynamic diffractive optics for enhanced femtosecond laser based cell transfection

High‐throughput laser‐mediated in situ cell purification with high purity and yield

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