Abstract
The introduction of nanoparticles to intact plant cells is promising as a transporting technique of a wide range of functional molecules. Among various molecular delivery methods, femtosecond laser photoinjection possesses target selectivity at a single cell level and is potentially applicable for many types of materials. However, for plant cells, the vacuoles’ turgor pressure and the thick cell wall limit the application of photoinjection to only small objects. In this work, we overcome these limitations by employing a single pulse irradiation from a femtosecond laser amplifier. After laser irradiation on intact tobacco BY-2 cells, 80 nm fluorescent nanoparticles dispersed in a cell culture medium were successfully injected into their cytoplasm. This breakthrough would lead to a vast utilization of nanoparticles containing functional molecules for single cell manipulation in plant physiological study and genetic engineering. Such an injection was observed even when the laser pulse was focused neither on the cell wall nor on the cell membrane, but beside the cells. With these results, we suggest pore formation on the cell membrane by instantaneous deformation induced by an intense femtosecond laser pulse as an injection mechanism of nanoparticles. Reported photomechanical effects of the amplified femtosecond laser on the permeability of the biological membrane would offer new perspectives in biophotonics.
Highlights
The introduction of nanoparticles to intact plant cells is a promising technique in agriculture and plant physiological study
Morphological changes in the tobacco BY-2 (TBY-2) cell structure after mannitol and enzyme treatment were confirmed by transmission images before fs laser photoinjection (Fig. 3)
After the mannitol addition to the culture medium, the cell vacuoles contracted in the hypertonic mannitol solution and the cell membrane was separated from the cell wall, namely, a plasmolyzed state [Fig. 3(b)]
Summary
The introduction of nanoparticles to intact plant cells is a promising technique in agriculture and plant physiological study. The introduction of nanoparticles into intact plant cells is generally managed either by diffusion through the plant cell wall or by active uptake mediated by cellular processes.. Our attention is directed to a laserassisted injection technique called photoinjection. In this method, molecules are injected through a pore formed by laser irradiation on a cell. Photoinjection possesses target selectivity at a single cell level with high throughput and it is applied to various materials.. Photoinjection possesses target selectivity at a single cell level with high throughput and it is applied to various materials.12–16 These features are significantly important to reveal spatiotemporally complex plant systems such as cell–cell communications. Photoinjection possesses target selectivity at a single cell level with high throughput and it is applied to various materials. These features are significantly important to reveal spatiotemporally complex plant systems such as cell–cell communications.
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