Low Stress Ion Conductance Microscopy of Sub-Cellular Stiffness

Abstract

Quantifying forces inherent to ion conductance microscopy enables it to map the stiffness of sub-cellular structures, even if very soft.

Visualising nanoscale restructuring of a cellular membrane triggered by polyelectrolyte microcapsules

“…S3 † ). Due to very low stress imposed by the nanopipette (typically in the range of 1–100 Pa) 19 the expected elastic modulus of capsules adhered to bare glass is beyond the measurement range of the method used here. Stress of 100 Pa would cause the capsule to compress by less than 5 pm (assuming a capsule with a diameter of 5 μm and an equivalent elastic modulus of at least 100 MPa), three orders of magnitude below the practical resolution limit of SICM.…”

“…To investigate the mechanical properties of capsule internalisation we used the recently implemented method for the measurement of elastic modulus. 19 Thanks to the low stress and minimally invasive character of the imaging using nanopipettes we were able to map changes of mechanical properties without apparent disruption of the internalisation process. The area of the cell membrane above the cell nuclei and around the nuclei displayed a mean elastic modulus of 4205 Pa ± 1284 ( n = 9) and 1310 Pa ± 559 ( n = 27), respectively.…”

“…The stress on the sample is caused by the intrinsic colloidal pressure between the cell surface and the glass tip at higher set points (>1%) which varies with the size of the tip and set point value, and is kept constant during a scan. 19 The elastic modulus of the sample is then calculated simply as a ratio of the applied stress to the observed strain between the two set points at each imaging point using custom built image processing software, SICM Image Viewer, using the algorithm developed by Clarke et al . 19 The elasticity maps have the same image resolution (effective pixel size) as topographical images, which was typically 320 nm for a 30 μm × 30 μm scan and 160 nm for a 15 μm × 15 μm scan.…”

Rapid formation of human immunodeficiency virus-like particles

“…This can be achieved by deforming the PM either by using hydrostatic pressure to fire a jet of liquid from the tip of the pipette (30,31) or by using intrinsic repulsion forces that appear in close-range interactions between the negatively charged glass of the pipette and negatively charged lipid membrane. These interactions occur at high set points (e.g., 2 to 3%) compared to the set point typically used for imaging, that is, 0.4 to 0.5% of ion current drop (32,33). In our experiments we used the second method as it allowed us to use smaller-diameter pipettes and thereby achieve higher spatial resolution when mapping cell stiffness.…”

“…It has been shown that at 2% set point a 110-nm-aperture pipette exerts sufficient force to push the PM 100 to 200 nm, not enough to deform the underlying cytoskeleton. A 3% set point is necessary to create a higher force required to measure the stiffness of the underlying actin cortex (32). Thus, low-force stiffness mapping can be used to report PM tension.…”