Jan 11, 2005
Nanoneedle gets into cells
Scientists at the Research Institute for Cell Engineering at Japan's National Institute of Advanced Industrial Science and Technology (AIST) and Tokyo University of Agriculture and Technology have used nanoneedles attached to an atomic force microscope (AFM) to penetrate the nucleus of living cells.
The researchers reckon that they could use the needles to deliver molecules such as nucleic acids, proteins or other chemicals to the nucleus, or maybe even to carry out cell surgery.
Standard sharp AFM tips were not suitable to act as needles as they were not long enough for the cells under test, which were more than 3 microns high. So the scientists etched the needles from pyramidal silicon AFM tips using focused ion beam etching.
"We have spent a long time seeking the appropriate needle material for insertion into the cell," researcher Chikashi Nakamura told nanotechweb.org. "At first, we tried carbon nanotube probes, but there was a problem with mechanical toughness. Finally, we settled on using an etched-silicon AFM tip sharpened to needle shape. Now the thinnest needles we can make have a diameter of 100 nm."
The nanoneedles the scientists tested were 200-300 nm in diameter and 6-8 microns long. The team tested two tip shapes - a flat-ended cylinder and a cone - on human epidermal melanocytes. The two types of needle produced different force-distance curves in the AFM.
"The cylindrical needle shape is important for invasiveness and success of insertion into the cell," said Nakamura.
The team also tested nanoneedles fashioned from an AFM probe with a tetrahedral tip on human embryonic kidney cells that expressed a red-fluorescent protein. The researchers coated the needles with a fluorescent dye and examined their position in the cell using laser scanning confocal microscopy. These cells were 10-20 microns high.
The experiment showed that the nanoneedles penetrated both the cellular and nuclear membranes and reached the nucleus of the cells. It also indicated that a force drop or force relaxation measured by the AFM was related to the successful insertion of the needle through the cell membrane.
The scientists say this is the first time that solid material was inserted into the nucleus of such a small living cell with highly accurate positioning.
According to the scientists, the nanoneedle technique has a number of advantages over microcapillary-based techniques: the size of the cells is not important as the needle displacement is accurate; the technique can estimate the position of the needle by monitoring the exerting force; and it can use thinner needles as there is no need for optical observation - this means that more of the cells survive.
"We propose that, in the future, differentiated cells prepared by multiple steps of manipulations using nanoneedles from stem cells can be used for cell therapy in the medical field," said Nakamura.
Now the team is trying to transfer substances such as nucleic acids, proteins and chemicals to cells. "Soon we will publish our result of gene transfer with high efficiency using a nanoneedle," said Nakamura.
The researchers reported their work in Nano Letters.
About the author
Liz Kalaugher is editor of nanotechweb.org.