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Pflügers Archiv - European Journal of Physiology

, Volume 464, Issue 3, pp 307–316 | Cite as

Measuring the elastic properties of living cells through the analysis of current–displacement curves in scanning ion conductance microscopy

  • Mario Pellegrino
  • Monica Pellegrini
  • Paolo Orsini
  • Elisabetta Tognoni
  • Cesare Ascoli
  • Paolo Baschieri
  • Franco Dinelli
Signaling and cell physiology

Abstract

Knowledge of mechanical properties of living cells is essential to understand their physiological and pathological conditions. To measure local cellular elasticity, scanning probe techniques have been increasingly employed. In particular, non-contact scanning ion conductance microscopy (SICM) has been used for this purpose; thanks to the application of a hydrostatic pressure via the SICM pipette. However, the measurement of sample deformations induced by weak pressures at a short distance has not yet been carried out. A direct quantification of the applied pressure has not been also achieved up to now. These two issues are highly relevant, especially when one addresses the investigation of thin cell regions. In this paper, we present an approach to solve these problems based on the use of a setup integrating SICM, atomic force microscopy, and optical microscopy. In particular, we describe how we can directly image the pipette aperture in situ. Additionally, we can measure the force induced by a constant hydrostatic pressure applied via the pipette over the entire probe–sample distance range from a remote point to contact. Then, we demonstrate that the sample deformation induced by an external pressure applied to the pipette can be indirectly and reliably evaluated from the analysis of the current–displacement curves. This method allows us to measure the linear relationship between indentation and applied pressure on uniformly deformable elastomers of known Young’s modulus. Finally, we apply the method to murine fibroblasts and we show that it is sensitive to local and temporally induced variations of the cell surface elasticity.

Keywords

Scanning ion conductance microscopy Atomic force microscopy Cell elasticity 

Abbreviations

SS

saline solution

R

electrical resistance of the pipette

ri

internal radius of the pipette aperture

ΔP

pressure applied to the pipette in addition to atmospheric pressure

z

probe displacement along the vertical axis (positive going down)

I

ion current flowing through the pipette

I0

maximum current value, measured far from the sample surface

I/I0

ratio of the actual current value to the maximum value

I/z

curve of the current measured versus pipette displacement

s

distance between pipette aperture and target surface

F

force exerted by the solution flowing out from the pipette aperture when ΔP ≠ 0

\( {F_{{\max }}} = \Delta P\;\pi \;r_i^2 \)

theoretical value of the force exerted by the flux at the pipette aperture

F/z

operating mode for AFM, yielding force versus displacement curves

d

deflection of the cantilever, measured via AFM

d/z

curve of the deflection measured versus pipette displacement

kC

nominal spring constant (0.01 N/m.)

dmax = Fmax / kC

theoretical deflection value expected at the contact between pipette and cantilever

dct

deflection value measured at the establishment of the contact

d/I

curve of relative deflection (d/d ct ), versus current ratio (I/I 0 )

δ

target deformation calculated by comparing two approaching curves obtained at different pressure, up to contact

δ1,2

target deformation calculated between two fixed I/I 0 values

δct.

target deformation calculated at contact

δ/I

curve of relative deformation (δ/δ ct ) versus current ratio (I/I 0 )

Notes

Acknowledgments

The authors wish to thank Francesco Montanari for his expert technical assistance. This work has been partially supported by funds of the Scuola Normale Superiore of Pisa.

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Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Mario Pellegrino
    • 1
  • Monica Pellegrini
    • 2
  • Paolo Orsini
    • 1
  • Elisabetta Tognoni
    • 3
  • Cesare Ascoli
    • 3
  • Paolo Baschieri
    • 3
  • Franco Dinelli
    • 3
  1. 1.Dipartimento di Scienze FisiologicheUniversità di PisaPisaItaly
  2. 2.Scuola Normale Superiore di PisaPisaItaly
  3. 3.Istituto Nazionale di Ottica del CNRPisaItaly

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