Femtosecond Laser-Assisted Optoporation for Drug and Gene Delivery into Single Mammalian Cells

Abstract

In this work, focused near-infrared (NIR) femtosecond laser pulses were used to transiently perforate the cellular membrane of targeted human embryonic kidney (HEK) cells and the uptake of extrinsic molecules into the targeted cells was observed. Various cellular responses to the laser treatments were closely analyzed to optimize several experimental parameters such as laser power, exposure time and location of laser irradiation using a membrane impermeable fluorescent dye. The optimized parameters were used to investigate the entry of a plasmid DNA encoding green fluorescent protein (GFP) into the target cells. Since laser beam with higher-than-threshold energy level will disintegrate cells, we used Matlab simulations to characterize the laser irradiance and free electron distribution caused by the femtosecond-optoporation process. The simulation results showed that the free electron distribution is much narrower than the laser irradiance, which implies that the transient perforation can even be smaller than the size of the laser focal volume. Femtosecond laser-assisted optoporation when combined with lab-on-a-chip devices can be useful in single cell-based high-throughput screening.

Optical delivery of multiple opsin-encoding genes leads to targeted expression and white-light activation

“…Recently, there has been considerable interest in the optical transfection of cells using ultrafast pulsed light [15][16][17][18][19][20][21][22][23] because this approach offers selective targeting and higher efficiency and viability (.90% reported in vitro 22 ) than other methods. Furthermore, femtosecond (fs) near-infrared (NIR) laser-based transfection has been shown to be safe, resulting in high efficiency and high survival rates (93%) of optoporated embryos during development 20 , as well as suitable for in vivo gene delivery 24 .…”

Advances in Applications of Dendritic Compounds

“…11 Apart from the above, click chemistry including Diels-Alder reactions and Thiolyne reactions have been employed to synthesize polyphenylene dendrimers. [12][13][14][15][16][17][18] Since then, based on the previous synthetic methods, in order to endow more features to new-style macromolecules, some researchers also synthesized diverse metal (non-metal) as well as dual-core and multi-core dendritic compounds with some functional groups. [19][20][21][22] …”

Physical energy for drug delivery; poration, concentration and activation

“…Biophysical energy such as electric fields, magnetic fields, ultrasound and mechanical forces, light and temperature gradients can act as enhancers for drug delivery. Technique-specific terms have been associated with each form of physical energy such as electroporation [37, 38] for electric current, iontophoresis for electric potential, magnetoporation [39, 40] for magnetic field, sonoporation [41, 42], mechanoporation [43] and phonophoresis [44, 45] for ultrasound, optoporation [46, 47] for pulsed light and thermoporation [48, 49] for temperature, respectively (Figure 1). Table 1 summarizes all of the different types of physical energy used in this review for porating the cells.…”