Uniform Transfection: Shock Wave Generation in Laser Ablation and Microcontact Printing
01 Dec 2018-pp 1-5
TL;DR: This research explores the possibility of using $\mu$ CP (using cell adhesive and cell blocking agents) for printing massive number of cells on a substrate and using optoporation methods (nano-second pulsed radiation) to perforate cell membranes to perform intracellular delivery.
Abstract: With the increasing need of regenerative medicines, cell therapy, and biopharmaceutical drug, it is necessary to develop a method for massively parallel delivery and uniform transfection. This research explores the possibility of using $\mu$ CP (using cell adhesive and cell blocking agents) for printing massive number of cells on a substrate and using optoporation methods (nano-second pulsed radiation) to perforate cell membranes. With combining these two methods, a stand-alone micro-device capable of performing localized intracellular delivery will be developed. In this paper, we confirmed the generation of shock wave upon optoporation, tried intracellular delivery using Calcein, develop the stamp for microcontact printing and performed the printing.
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TL;DR: A highly efficient gene delivery of over 70% was achieved in human MSCs, which compared very favorably with other major nonviral gene delivery methods (lipofection approximately 50%, microinjection approximately 10 %).
Abstract: We describe a low-invasive gene delivery method that uses an etched atomic force microscopy (AFM) tip or nanoneedle that can be inserted into a cell nucleus without causing cellular damage. The nanoneedle is 200 nm in diameter and 6 μm in length and is operated using an AFM system. The probabilities of insertion of the nanoneedle into human mesenchymal stem cells (MSCs) and human embryonic kidney cells (HEK293) were higher than those of typical microinjection capillaries. A plasmid containing the green fluorescent protein (GFP) gene was adsorbed on a poly- l -lysine–modified nanoneedle surface, which was then inserted into primary cultured single human MSCs. A highly efficient gene delivery of over 70% was achieved in human MSCs, which compared very favorably with other major nonviral gene delivery methods (lipofection ~50%, microinjection ~10 %). The single cells expressing GFP were collected and the amount of delivered DNA in each cell was analyzed. The highest rate of expressed GFP per delivered DNA was achieved using the nanoneedle, because the nanoneedle could be inserted into the nucleus directly without causing significant cell damage.
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