A method that uses a femtosecond laser to poke a hole in the surface of a single cell and gently tug a piece of DNA through it with optical tweezers offers greater control for gene therapy and genetic engineering. Researchers at the School of Mechatronics at Gwangju Institute of Science and Technology used optical tweezers to grab a plasmid-coated particle and then transport it to the surface of the cell. Guided by the trapped particle, they used an ultrashort laser pulse to create a tiny pore in the cell membrane. While another laser beam detected the exact location of the membrane, they pushed the particle through the pore with the tweezers and dropped it easily into the cell. Plasmid-coated particles can be optically manipulated and inserted into a cell through a small pore made by a short-pulsed laser. Plasmids produce a green fluorescent protein once inside the cell. (Drawing is not to scale.) Courtesy of Gwangju Institute of Science and Technology. To determine whether the method had succeeded, the researchers inserted plasmids carrying a gene that codes for a green fluorescent protein. Once inside the cell, the gene became active, and the cell’s machinery began producing the protein. The researchers could then detect the green glow using a fluorescence microscope. They found that approximately one in six of the cells they studied became transfected. This rate is lower than that recorded for some other methods, but those are less precise and involve many cells at a time. “What is magical is that all this happens for one cell,” said associate professor Yong-Gu Lee. “Until today, gene transfection has been performed on a large quantity of agglomerate cells, and the outcome has been observed as a statistical average, and no observations have been made on individual cells.” (a) A laser scanning microscope image of a cancer cell used in the experiment. The green circles show plasmid-coated particles that have been optically tweezed and inserted into the cell. (b) The same cell viewed with a fluorescence microscope. The DNA material inserted into the cell through the transfection process carries a gene that codes for a green fluorescent protein. Here, the cell’s green glow means the transfection process was successful. (c) Image (b) superimposed on image (a). Courtesy of Biomedical Optics Express. Lee hopes the work will allow other researchers to investigate the effects of transfection on individual cells, not just on large populations. With the new technique, “you can put one gene into one cell, another gene into another cell, and none into a third,” he said. “So you can study exactly how it works.” The study was reported in The Optical Society’s Biomedical Optics Express (doi: 10.1364/boe.4.001533).