Atomic force microscopy (AFM) allows the acquisition of high-resolution images and the measurement of mechanical properties of living cells under physiological conditions. AFM cantilevers with blunted pyramidal tips are commonly used to obtain images of living cells. Measurement of mechanical properties with these tips requires a contact model that takes into account their blunted geometry. The aim of this work was to develop a contact model of a blunted pyramidal tip and to assess the suitability of pyramidal tips for probing mechanical properties of soft gels and living cells. We developed a contact model of a blunted pyramidal tip indenting an elastic half-space. We measured Young's modulus (E) and the complex shear modulus (G*= G' +i G" ) of agarose gels and A549 alveolar epithelial cells with pyramidal tips and compared them with those obtained with spherical tips. The gels exhibited an elastic behavior with almost coincident loading and unloading force curves and negligible values of G". E fell sharply with indentation up to approximately 300 nm , showing a linear regime for deeper indentations. A similar indentation dependence of E with twofold lower values at the linear regime was obtained with the spherical tip fitted with Hertz's model. The dependence of E on indentation in cells paralleled that found in gels. Cells exhibited viscoelastic behavior with G"/G' approximately 1/4 . Pyramidal tips commonly used for AFM imaging are suitable for probing mechanical properties of soft gels and living cells.

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Probing mechanical properties of living cells by atomic force microscopy with blunted pyramidal cantilever tips.

The contact of an indenter of arbitrary shape on an elastically anisotropic half space is considered. It is demonstrated in a theorem that the solution of the contact problem is the one that maximizes the load on the indenter for a given indentation depth. The theorem can be used to derive the best approximate solution in the Rayleigh–Ritz sense if the contact area is a priori assumed to have a certain shape. This approach is used to analyze the contact of a sphere and an axisymmetric cone on an anisotropic half space. The contact area is assumed to be elliptical, which is exact for the sphere and an approximation for the cone. It is further shown that the contact area is exactly elliptical even for conical indenters when a limited class of Green's functions is considered. If only the first term of the surface Green's function Fourier expansion is retained in the solution of the axisymmetric contact problem, a simpler solution is obtained, referred to as the equivalent isotropic solution. For most anisotropic materials, the contact stiffness determined using this approach is very close to the value obtained for both conical and spherical indenters by means of the theorem. Therefore, it is suggested that the equivalent isotropic solution provides a quick and efficient estimate for quantities such as the elastic compliance or stiffness of the contact. The “equivalent indentation modulus”, which depends on material and orientation, is computed for sapphire and diamond single crystals.

http://www-personal.umich.edu/%7Ejbarber/vlassak.pdf

The indentation modulus of elastically anisotropic materials for indenters of arbitrary shape

We present a procedure that allows a reliable determination of the elastic (Young's) modulus of soft samples, including living cells, by atomic force microscopy (AFM). The standardized nanomechanical AFM procedure (SNAP) ensures the precise adjustment of the AFM optical lever system, a prerequisite for all kinds of force spectroscopy methods, to obtain reliable values independent of the instrument, laboratory and operator. Measurements of soft hydrogel samples with a well-defined elastic modulus using different AFMs revealed that the uncertainties in the determination of the deflection sensitivity and subsequently cantilever's spring constant were the main sources of error. SNAP eliminates those errors by calculating the correct deflection sensitivity based on spring constants determined with a vibrometer. The procedure was validated within a large network of European laboratories by measuring the elastic properties of gels and living cells, showing that its application reduces the variability in elastic moduli of hydrogels down to 1%, and increased the consistency of living cells elasticity measurements by a factor of two. The high reproducibility of elasticity measurements provided by SNAP could improve significantly the applicability of cell mechanics as a quantitative marker to discriminate between cell types and conditions.

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Standardized Nanomechanical Atomic Force Microscopy Procedure (SNAP) for Measuring Soft and Biological Samples.

In this paper, the responses of nanoindentation on bulk silicon were investigated using finite element analysis. A two-dimensional finite element model under the assumption of axisymmetry was successfully validated by the experimental load–displacement curve. Four factors: coefficient of friction, indentation depth, tip rounding and indenter geometry were investigated to characterize the induced responses of bulk silicon via load–displacement curve, indentation surface profile at the maximum loading depth, residual surface profile after unloading, plastic energy, elastic energy, Young's modulus, hardness, and elastic recovery. Coefficients of friction were found to be insignificant, but the von Mises stress distributions after unloading between frictionless and frictional surfaces, indentation depth, tip rounding, and indenter geometry all showed having a distinct effect on stress, plastic energy, elastic energy, Young's modulus, hardness, elastic recovery and surface profile. The degree of pile-up affecting the investigated factors is discussed as well.

A numerical study of factors affecting the characterization of nanoindentation on silicon

A general three-dimensional contact, between elastically similar half-spaces, is considered. With a fixed normal load, we consider a pure relative tangential translation between the two bodies. We show that, for the case of negligible Poisson's ratio, an exact solution is given by a single component of shearing traction, in the direction of loading. It is well known that, for full sliding conditions, the tangential force must be applied through the center of the pressure distribution. Instead, for a full stick case the tangential force must be applied through the center of the pressure distribution under a rigid flat indenter whose planform is the contact area of the problem under consideration. Finally, for finite friction a partial slip regime has to be introduced. It is shown that this problem corresponds to a difference between the actual normal contact problem, and a corrective problem corresponding to a lower load, but with same rotation of the actual normal indentation. Therefore for a pure translation to occur in the partial slip regime, the point of application of the tangential load must follow the center of the difference' pressure. The latter also provides a complete solution of the partial slip problem. In particular, the general solution in quadrature is given for the axisymmetric case, where it is also possible to take into account of the effect of Poisson's ratio, as shown in the Appendix.

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D.A. Billings

Papers

An approximate solution for the contact area and elastic compliance of a smooth punch of arbitrary shape