Nanoscale Operation of a Living Cell Using an Atomic Force Microscope with a Nanoneedle

Abstract

We have developed a tool for performing surgical operations on living cells at nanoscale resolution using atomic force microscopy (AFM) and a modified AFM tip. The AFM tips are sharpened to ultrathin needles of 200-300 nm in diameter using focused ion beam etching. Force-distance curves obtained by AFM using the needles indicated that the needles penetrated the cell membrane following indentation to a depth of 1-2 microm. The force increase during the indentation process was found to be consistent with application of the Hertz model. A three-dimensional image generated by laser scanning confocal microscopy directly revealed that the needle penetrated both the cellular and nuclear membranes to reach the nucleus. This technique enables the extended application of AFM to analyses and surgery of living cells.

An historical perspective on cell mechanics

“…c The modern AFM, integrated with an inverted laser scanning confocal microscope to allow simultaneous mechanical perturbations and measurements to be performed while imaging cellular structures in three dimensions needles were used to push and penetrate into organelles of living cells [135]. Interestingly, in 2005, a "nanoscale operation" was described in which an AFM tip, modified with a nanoneedle, was employed to push and penetrate into the nucleus of living cells [136]. Although separated by about three quarters of a century, both reports describe the penetration and deformation of the cell nucleus using very similar approaches (Fig.…”

“…b Much later, modern techniques using AFM as seen in the phase-contrast micrograph. These similar methods of "nano-indentation" have also been described as "nano-operations" [136] control the heart by stimulating it correctly. Although the measurement of accurate mechanical parameters is of extreme importance, the idea of controlling and altering biological pathways is equally enticing.…”

ATP-mediated Release of a DNA-binding Protein from a Silicon Nanoneedle Array

“…HeLa cells expressing red fluorescent protein, DsRed2-NES, whose plasmid was constructed in our laboratory 19 were seeded (10 5 cells) on a collagen-coated 12] glass base dish (IWAKI) and cultured in high glucose DMEM supplemented with 10% fetal bovine serum (FBS). The medium were changed to DMEM/F-12, which does not contain either phenol red or FBS, prior to insertion of the nanoneedles into the cell.…”

Intracellular manipulation of chromatin using magnetic nanoparticles

“…Recently, a modified AFM using nanoneedles was developed (Obataya et al 2005a). These nanoneedles can be used to penetrate the cell membrane to manipulate internal cellular structures (Obataya et al 2005b). AFM allows careful positioning of the nanoneedle and simultaneous measurement of the applied forces.…”